Homocysteine induces expressions of adhesive molecules on leukocytes in whole blood

Objective To observe whether homocysteine can directly alter the expressions of CD11b, CD1 8, CD14 and L-selectin on neutrophils, monocytes, and lymphocytes in whole bloo d from healthy human subjects. Methods Leukocyte surface adhesive molecule expressions were analyzed by immunofluoresce nce flow cytometry.Results Homocysteine at the lowest concentration (20 μmol/L) significantly increased s urface adhesive molecule expressions of CD11b and CD18 on each cell type and CD1 4 on monocytes and neutrophils in whole blood.These effects were increased at homocysteine concentrations of 200 and 400 μmol/L, but at concentrations 1 mmol/L, CD11b/CD18 and CD14 expressions on all types of leukocytes were decrea sed.L-selectin expression was slightly decreased on all cell types in whole blood by homocysteine.Conclusion Homocysteine alters leukocyte expressions of CD11b/CD18, CD14 and L-selectin on leukocytes, which may be involved into homocysteine-induced leukocyte adhesion and migration.     

Inflammation induced-endothelial cells release angiogenesis associated-microRNAs into circulation by microparticles

Background Endothelial cells derived microRNAs can be detected in plasma and serum and there is evidence that inflammatory disease states may affect the levels of circulating microRNAs.However,there is no direct proof that inflammation induces endothelial cells to release microRNAs into circulation.This study aimed to explore whether inflammation could induce endothelial cells to release microRNAs into circulation and to investigate whether these released microRNAs derived from endothelial cells were transported in microparticles.Methods Microparticles were isolated from human atherosclerotic plaques with an active inflammatory phenotype and normal vascular tissue.Flow cytometry and real-time PCR were used to detect the levels of microparticles and microRNAs.Human umbilical vein endothelial cells （HUVEC） was treated with tumour necrosis factor α （TNF-α,10 ng/ml） for 24 hours,and then HUVEC and the culture medium were respectively collected.Results By comparing microparticles isolated from human atherosclerotic plaques with an active inflammatory phenotype （n=9） and those from normal vascular tissues （n-9）,we found levels of annexin V＋ microparticles and annexin V＋ CD144＋ microparticles were significantly increased in plaques and angiogenesis associated microRNAs （106b,25,92a and 21） were also significantly increased in microparticles from plaques.After exposure to TNF-α at a concentration of 10 ng/ml （TNF-α group,n=3） or DMEM （control group,n=3） for 24 hours,counts of microparticles and expressions of microRNAs 106b,25,92a and 21 in microparticles isolated from medium significantly increased.However,there were no differences in the intracellular levels of microRNAs 25,92a or 21 isolated from HUVEC between TNF-α group and control group,while microRNA 106b decreased in TNF-α group.Conclusion Inflammation could induce endothelial cells to release angiogenesis associated microRNAs into circulation,causing higher levels of circulating endothelial cells derived microRNAs in ather     

Changes of T cell subsets in peripheral blood of patients with Schistosomiasis japonica and their relation to interleukin-1.

T cell subsets in peripheral blood were phenotyped in 56 patients with different stages of Schistosomiasis japonica, including 17 with acute, 14 with chronic and 25 with advanced infection. The activity of interleukin-1 (IL-1) produced by peripheral blood mononuclear cells (PBMC) induced by lipopolysaccharide (LPS) in vitro was simultaneously detected in these three groups of patients. It was found that the percentages of CD3+ (total T cell), CD4+ (helper/inducer T cell) and CD8+ (suppressor/cytotoxic T cell) T cell and the level of IL-1 were significantly increased in the group of acute Schistosomiasis japonica. In the groups of chronic and advanced Schistosomiasis japonica, the proportion of CD3+ T cell, the ratio of CD3+/CD4+ and the level of IL-1 were remarkably reduced, and the percentage of CD8+ T cell was increased. The rate of CD4+ T cell was obviously decreased in cases patients with advanced Schistosomiasis japonica. The percentage of CD4+ T cell was positively correlated to the level of IL-1 in the three groups of patients. These results indicate that T cell subsets and IL-1 may play an important role in the immunoregulation of Schistosomiasis japonica.

     

Cardiotrophin-1 induces intercellular adhesion molecule-1 expression by nuclear factor κB activation in human umbilical vein endothelial cells

Background In addition to elevated concentrations of cytokines, patients with congestive endothelial dysfunction and increased plasma concentrations of adhesion molecules heart failure （CHF） show ke intercellular adhesion molecule-1 （ICAM-1）. Furthermore, the concentration of cardiotrophin-1 （CT-1） - a cytokine of the interleukin-6 superfamily - is increased in CHF. We tested the hypothesis whether CT-1 is able to induce ICAM-1 in human umbilical vein endothelial cells （HUVEC）. Furthermore we examined the signalling mechanisms of CT-1 mediated ICAM-1 expression. Methods Confluent layers of HUVEC were incubated with increasing concentrations of CT-1 （5 to 100 ng/ml） for different periods. ICAM-1 mRNA was determined by real-time polymerase chain reaction （PCR） and ICAM-1 surface expression by fluorescence-activated cell sorter （FACS） analysis and soluble ICAM-1 （slCAM-1） in the culture supernatant by enzyme linked immunosorbent assay （ELISA）. To clarify the signalling pathway of CT-1 induced ICAM-1 expression we used various inhibitors of possible signal transducing molecules, electromobility shift assay （EMSA） and Western blot analysis. Results CT-1 induced ICAM-1 mRNA （1.8±0.8 fold increase compared to unstimulated cells after 6 hours） and protein （1.4±0.2 fold increase compared to unstimulated cells after 48 hours） in HUVEC in a time- and concentration-dependent manner. EMSA experiments show that CT-1 causes nuclear factor （NF） κB activation. Because parthenolide could inhibit CT-1 induced ICAM-1 expression NFκB activation is required in this pathway. CT-1 did not activate extracellular signal regulated kinases （ERK）, c-Jun N-terminal kinase （JNK） and p38. Conclusion CT-1 is able to induce ICAM-1 in endothelial cells by NFκB activation. These results may explain in part elevated ICAM-1 concentrations in patients with CHF and endothelial dysfunction.     

Role of pigment epithelium-derived factor on proliferation and migration of choroidal capillary endothelium induced by vascular endothelial growth factor in vitro

Background Pigment epithelium-derived factor （PEDF） is expressed in several normal organs and identified as an inhibitor of neovascularization. In the present study, we investigated the effect of PEDF in an in vitro model of ocular choroidal neovascularization. Methods Microdissection was used to isolate the human choroidal endothelial cells （CECs）, followed by the use of superparamagnetic beads （Dynabeads） coated with the CD31 antibody, which selectively binds to the endothelial cell surface. The mitogenic and motogenic effects of vascular endothelial growth factor （VEGF） on cultured choroidal capillary endothelial cells were examined in the presence or absence of PEDF （1, 10, 100, and 1000 ng/ml） using cell counts and migration assays. Results Cells bound to the beads were isolated using a magnetic particle concentrator and they were successfully cultured and characterized to be endothelial cells that possessed greater than 95% immunoreactivity to von Willebrand factor. PEDF suppressed the proliferation and migration of VEGF-induced choroidal capillary endothelial cells. However, the concentration of PEDF which we used has little effect on normal CECs. Conclusions PEDF played an important role on the growth and migration of VEGF-stimulated choroidal endothelial cell These findings suggest that PEDF may be an effective approach to the treatment of choroidal neovascular disorders.     

Vascular endothelial growth inhibitor affects the invasion, apoptosis and vascularisation in breast cancer cell line MDA-MB-231

Background Breast cancer is one of the most common malignant female diseases worldwide.It is a significant threat to every woman＇s health.Vascular endothelial growth inhibitor （VEGI） is known to be abundant in endothelial cells.According to previous literature,overexpression of VEGI has been shown to inhibit tumor neovascularisation and progression in cellular and animal models,but there has been limited research on the significance of VEGI in the breast cancer.Methods In our study,cell lines MDA-MB-231 were first constructed in which VEGI mediated by lentivirus over-expressed.The effects of VEGI over-expression on MDA-MB-231 cells were investigated both in vitro and in vivo.The expression of VEGI in the MDA-MB-231 cells after infection of lentivirus was analyzed using real-time PCR and Western blotting.The effect of the biological characteristics of MDA-MB-231 cells was assessed by growth,invasion,adhesion,and migration assay with subcutaneous tumor-bearing nude mice models.Then the growth curves of the subcutaneous tumors were studied.Expressions of VEGI,CD31 and CD34 in the tumors were analyzed by immunohistochemistry and apoptosis was detected by flow cytometry and immunohistochemistry.Results Infection of MDA-MB-231 cells within the lentivirus resulted in approximately a 1 000-fold increase in the expression of VEGI.As can be seen in the invasion,adhesion and migration assay,the over-expression of VEGI can inhibit the ability of MDA-MB-231 cells during migration,adhesion and invasion.The volume of the subcutaneous tumor in the over-expression group was distinctly and significantly less than that of the control groups.Immunohistochemistry analysis of the tumor biopsies cleady showed the expression of VEGI in the over-expression group increased while CD31 and CD34 decreased significantly.In vitro and in vivo,the early apoptosis rate and the apoptosis index were increased within the VEGI over-expression group as compared with the control group.Conclusions Taken together,recombinant lentivirus that were successfully constructed,demonstrated up-regulated VEGI gene expression in breast cancer cells.Lentivirus-mediated over-expression of VEGI weakened the ability of the breast cancer cell migration,adhesion and invasion.Over-expression of VEGI diminished the tumorigenic capacity of breast cancer cells in vivo.Up-regulation of VEGI gene expression however inhibited breast cancer MDA-MB-231 cell in the early apoptosis.     

维生素E对肿瘤坏死因子损伤 主动脉内皮细胞的防治作用

Objective　To explore the possible mechanism of vitamin E against atherosclerosis through its preventive effect on tumor necrosis factor (TNF) inducing endothelial cell injury.Methods　Bovine aortic endothelial cells were incubated in culture media with TNF-alpha (400 U/ml) and vitamin E (12.5 microM, 25.0 microM and 50.0 microM respectively) for 72 h.The morphology, proliferation, apoptosis and adherence of endothelial cells were investigated with phase-contrast microscope, MTT method, flow cytometry analysis and cell count respectively.Results　Compared with TNF group, the phenomena of endothelial cell deformity and shedding were slighter, the number of adhered HL60 monocyte cells and G₁-phase cells were less, and the inhibitory rate and the apoptotic peaks were significantly lower in vitamin E groups.Conclusion　The results suggested that vitamin E has definite protective effects on injury induced by TNF as shown in morphology, proliferation and apoptosis of endothelial cells in vitro.     

同型半胱氨酸增高血白细胞表面粘附分子的表达

目的　观察同型半胱氨酸是否改变体外健康人血单核细胞、中性粒细胞及淋巴细胞表面粘附分子CD11b ,CD18,CD14和L 选择素 (L selectin)的表达。方法　细胞表面粘附分子CD11b ,CD18,CD14和L selectin的测定采用直接免疫荧光染色 ,2 4小时内用流式细胞仪测定。结果　同型半胱氨酸在低浓度 (2 0 μmol/L)时明显增加CD11b和CD18在各种白细胞表面的表达 ,同时也增加了CD14在单核及中性粒细胞的表达。这种作用随同型半胱氨酸浓度升高而增强 ,但当浓度升至≥ 1mmol/L时 ,各种粘附分子的表达反而降低。同型半胱氨酸使各种细胞类型表面L selectin降低。结论　同型半胱氨酸可改变白细胞表面粘附分子的表达 ,可能在体内通过此途经导致白细胞粘附并游出于血管内皮。     

脑血栓形成患者白细胞CD11a,CD11b的表达变化研究

This study sought to understand the mechanism for the increased adhesion of leucocytes and endothelial cells in ischemic stroke. 20 patients with acute cerebral thrombosis and 20 healthy subjects as controls for expression of CD11a and CD11b (adhesion molecules on surface of leucocytes) were tested in vitro by flow cytometry (FCM) method. The results showed that compared with the control group, the patient group had significantly higher rates for expression of CD11a on monocytes, granulocytes and lymphocytes (P < 0.05). The CD11b expression in the patient group was positively elevated on monocytes and granulocytes (P < 0.05), but it was of lower positive rate on lymphocytes and no statistical difference was noted between the patient and control groups. These indicate that the expression of CD11a and CD11b on leucocytes increases in cerebral ischemic damage; thus adhesion of leucocytes and endothelial cells obviously increases. This change may aggravate post-ischemic delayed neuronal death.     

脑血栓形成患者白细胞CD_(11a)、CD_(11b)的表达变化研究

采用流式细胞术，用单克隆抗体定量测定了２０例急性脑血栓形成患者周围血中白细胞ＣＤ１１ａ、ＣＤ１１ｂ的阳性表达率，并以２０例健康人作对照。结果：脑血栓形成组血液ＣＤ１１ａ在单核细胞、粒细胞、淋巴细胞的阳性表达率均明显高于对照组（Ｐ＜０．０５）；ＣＤ１１ｂ在粒细胞和单核细胞的阳性表达率也明显高于对照组（Ｐ＜０．０５），但它在淋巴细胞上的表达两组间则无显著性差异。由此提示脑血栓形成时白细胞ＣＤ１１ａ、ＣＤ１１ｂ表达上调，白细胞与内皮细胞的粘附明显增强，可能会加重缺血后迟发性神经元死亡的病理过程。     

Study of the Mechanism of Essential Garlic Oil Inhibiting Interleukin1 Induced Monocyte Adhesion to Endothelial Cells

Ｇａｒｌｉｃｈａｓｂｅｅｎｕｓｅｄａｓａｎｅｆｆｅｃｔｉｖｅｍｅｄｉｃｉｎｅｆｏｒｃｅｎｔｕｒｉｅｓ．Ｉｎｒｅｃｅｎｔｙｅａｒｓ,ｇａｒｌｉｃｈａｓａｔｔｒａｃｔｅｄｍｏｒｅａｔｔｅｎｔｉｏｎｄｕｅｔｏｉｔｓｐｒｏｔｅｃ ｔｉｖｅｅｆｆｅｃｔｓａｇａｉｎｓｔｃａｒｄｉｏｖａｓｃｕｌａｒｄｉｓｅａｓｅｓ．Ｖａｓｃｕｌａｒｃｅｌｌａｄｈｅｓｉｖｅｍｏｌｅｃｕｌｅ 1 （ＶＣＡＭ 1,ＣＤ1 0 6）ｉｓａｍｅｍｂｅｒｏｆｉｍｍｕｎｏｇｌｏｂｕｌｉｎｓｕｐｅｒｆａｍｉｌｙ．Ｔｈｅａｄｈｅｓｉｖｅｍｏｌｅｃｕｌｅ…     

烧伤后CD11b／CD18介导的中性多形核白细胞粘附对肺微血管内皮？…

目的 观察烧伤后中性多形核白细胞（ＰＭＮ）对肺微血管内皮细胞单层通透性的影响,以及ＰＭＮ粘附及其粘附分子ＣＤ１１ｂ／ＣＤ１８在该影响中的介导作用。方法 用培养血管内皮细胞单层模型,建立培养肺微血管内皮细胞（ＰＭＥＣ）单层通透性测定的方法,根据处理内皮细胞单层的不同成份,将实验分为７组,用含荧光素异硫氰酸酯－白蛋白的灌流液灌流后,测定液体滤过系数（ｋｆ）和渗透压反射系数（δ）。结果 烧伤后ＰＭＮ能使     

流场对细胞黏附及其后内皮细胞VE-cadherin表达的影响

目的：探讨不同流场对THP-1单核细胞与ECV304内皮细胞黏附的影响及细胞黏附后内皮细胞VE-cadherin表达及分布。方法：不同流场作用单层内皮细胞及单核细胞6h,观察不同流场下单核细胞与内皮细胞黏附;用W estern蛋白印迹法检测黏附后VE-cadherin的表达,用免疫细胞化学方法在激光共聚焦显微镜下观察VE-cadherin蛋白在内皮细胞中的分布变化。结果：不同流场作用单层内皮细胞及单核细胞6 h,发现低剪切力层流组与高剪切力层流组黏附的单核细胞均数分别为59.8/视野及3/视野,低剪切力紊流组与低剪切力层流组黏附的单核细胞均数分别为125/视野及61/视野,低剪切力紊流下单核细胞黏附数高于低剪切力层流及高剪切力层流,且在低剪切力紊流组中有大量单核细胞成簇黏附于内皮细胞上;单核细胞与内皮细胞共育0、1、2、4 h后,VE-cadherin蛋白的表达量在各组细胞间无差异;0 h组VE-cadherin着色集中于内皮细胞周边即细胞与细胞之间,1h,2h,4h时VE-cadherin着色不再集中于细胞周边而是分散到整个内皮细胞膜。结论：低剪切力紊流环境下,单核细胞与内皮细胞的黏附增加且形成了较多的成簇黏附,细胞黏附后内皮细胞膜上VE-cadherin的分布随之改变。     

Effects of CD11/CD18 mediated PMN-EC adhesion on the endothelial cell damages in the early stage postburn in vitro

Activatedpolymorphonuclearneutrophils(PMNs)havebeenshowntoproducedamagesonvascularendothelialcells(ECs)throughthegener-ationofoxygen-derivedfreeradicalsandthere-leaseofproteases;andPMN-ECadhesionisessen-tialtomediateECdamages['j.TheintegrinCD11/CD18consistsof3heterodimericsubunits,eachofwhichcomprisesadistinctAchain(CDl1a,CDl1bandCD1lc)thatisnoncovalentlyassociatedwitlhacommonBchain(CD18).TheimportanceoftheCD1l/CD18integrininPMNadhesionandemigrationinsystemicvasculaturehasbeencom-…     

急性脑梗死患者白细胞粘附分子CD_(11a)、CD_(11b)的表达

目的:探讨缺血性脑血管病白细胞与内皮细胞粘附增强的机理。方法:采用流式细胞术,用单克隆抗体(McAb)定量测定了42例急性脑梗死患者周围血中白细胞CD11a、CD11b的阳性表达率并与40例健康人作对照。结果:急性脑梗死组血液CD11a在单核细胞、粒细胞、淋巴细胞的阳性表达率均明显高于对照组P<0.05CD11b在粒细胞和单核细胞的阳性表达率也明显高于对照组(P<0.05)但它在淋巴细胞上的表达两组间则无显著性差异。结论:急性脑梗死发生时白细胞CD11a、CD11b表达上调,白细胞与内皮细胞的粘附明显增强可能会加重缺血后迟发性神经元死亡的病理过程。     

ICAM—1在缺氧—再氧化引起的白细胞与内皮细胞粘附中的作用

目的探讨细胞间粘附分子（ＩＣＡＭ－１）是否介导缺氧－再氧化引起的中性粒细胞（ＰＭＮ）和血管内皮细胞（ＶＥＣ）的粘附反应。方法血管内皮细胞（ＶＥＣ）经缺氧再氧化（Ｈ／Ｒ）处理后,加入ＰＭＮ,用计数法检测粘附率,以细胞免疫化学及原位杂交法检测ＩＣＡＭ－１及ＩＣＡＭ－１ｍＲＮＡ表达。结果ＶＥＣ经Ｈ／Ｒ处理后,ＰＭＮ与其粘附率增高１倍（Ｐ＜０．０１）,ＩＣＡＭ－１单克隆抗体（ｍＡｂ）与ＣＤ１１ａ／ＣＤ１８ｍＡｂ可明显降低粘附率的增高,Ｈ／Ｒ能增加ＶＥＣ的ＩＣＡＭ－１及ＩＣＡＭ－１ｍＲＮＡ表达。结论ＩＣＡＭ－１介导Ｈ／Ｒ后的ＰＭＮ－ＶＥＣ间粘附反应。     

巴曲酶对急性缺血性脑卒中患者外周血中黏附分子表达的影响

目的：观察巴曲酶对急性缺血性脑卒中患者外周血白细胞黏附分子和血清中可溶性黏附分子表达水平的影响。方法：治疗组18例，予巴曲酶3d（20BU）并予其他常规治疗；对照组18例，除未予巴曲酶外，余用药同巴曲酶治疗组。采用流式细胞术和酶联免疫吸附法测定患者卒中后不同时间外周血中性粒细胞和单核细胞CD11b、CD18、CD62L、CD54的表达以及血清中可溶性ICAM－1、VCAM－1的水平。结果：与对照组比，巴曲酶治疗组卒中患者在发病24h后中性粒细胞CD11b的表达下降（P＜0.05），单核细胞CD11b、中性粒细胞和单核细胞CD18、CD62L、CD54的表达及血清中可溶性ICAM－1、VCAM－1的水平均无显著性变化。结论：巴曲酶对急性缺血性脑卒中患者外周血黏附分子的表达的影响有限，提示降低外周血黏附分子的表达可是巴曲酶治疗缺血性脑卒中的主要作用机制。