人肺泡上皮细胞A549中细胞间粘附分子1的siRNA构建探讨

【摘要】〓目的〓构建A549细胞中细胞间粘附分子 1（ICAM 1）的小干扰RNA,探讨ICAM 1特异性siRNA能否抑制LPS诱导A549细胞ICAM 1的表达。方法〓采用化学合成法构建A549细胞ICAM 1的小干扰RNA。LPS诱导A549细胞ICAM 1表达,用RT PCR和流式细胞仪分别从mRNA和蛋白水平检测ICAM 1的表达。使用阳离子脂质体Lipofectamin2000转染 ICAM 1siRNA到A549细胞,再用LPS刺激A549细胞,RT PCR和流式细胞仪分别检测RNA干扰A549细胞后ICAM 1的mRNA和蛋白表达。结果〓LPS可以诱导A549细胞ICAM 1的表达,但LPS浓度过大损伤细胞。10 ng/ml的LPS组和100 ng/ml的LPS组OD值相比,两组OD值之间差异有显著性(P ＜0001)。10 ng/ml的LPS作用A549细胞8 h左右,细胞无明显损伤;10 ng/ml LPS作用A549细胞1h后,ICAM 1 mRNA的表达上调（P ＜005）,而ICAM 1蛋白水平增加不明显;10 ng/ml LPS作用A549细胞4h后,ICAM 1 mRNA表达水平明显增加（P ＜005）,ICAM 1蛋白水平也增加明显。通过化学合成法构建ICAM 1的siRNA,筛选出最优siRNA。转染 ICAM 1siRNA后,LPS刺激A549细胞的ICAM 1的mRNA和蛋白表达水平增加不明显。结论〓通过化学合成法构建的ICAM 1siRNA,可以有效地干扰ICAM 1的表达,ICAM 1siRNA降低了LPS诱导的ICAM 1的表达。     

彩色多普勒测定新生儿颅内动脉血流正常值100例报告

目的： 研究新生儿脑血流的血流动力学参数。方法： 应用彩色多普勒检测正常新生儿脑动脉血流变化的规律性。结果： 同名双侧大脑动脉血流的收缩期峰流速 （ＰＳ）、舒张末流速 （ＥＤ）、阻力指数 （ＲＩ）、搏动指数（ＰＩ） 经统计学处理, 无显著性差异Ｐ＞ ０．０５。结论： 提出了国人正常新生儿ＡＣＡ, Ｍ ＣＡ, ＰＣＡ 血流参数的正常值范围。     

Modulation of ICAM‐1 tissue expression in patients with liver transplantation (LT) and acute rejection (AR) after glucocorticoid treatment

Abstract Acute rejection (AR) is a frequent complication following liver transplantation (LT). ICAM‐1 may be involved in its pathogenesis. High doses of glucocorticoids are the standard treatment in these patients. The aim of this study was to describe corticoid effects on ICAM‐1 tissue expression in liver biopsies of patients with LT and AR. The study included liver biopsies performed before and after treatment in 12 patients with LT and proven AR. In 10 patients AR was reversible and in 2, was steroid resistant. For immunohistochemistry, an indirect immunoperoxidase technique was used. Each histology section was semiquantitatively evaluated as follows: 0: <10% staining, 1: 10‐25%, 2: 25‐50%, 3: >50%. The control group comprised nine patients with LT and normal liver biopsies. In pre‐treatment liver biopsy samples, ICAM‐1 was markedly expressed on sinusoidal cells (2.41 ± 0.66), and there was also expression on periportal (0.66 ± 0.65) and perivenular hepatocytes (0.83 ± 0.57). By contrast, in the liver tissue from the control group, sinusoidal ICAM‐1 reactivity was significantly lower (0.88 ± 0.33; P < 0.05), and hepatocytes showed no reliable ICAM‐1 expression. After steroid treatment the intensity of ICAM‐1 decreased significantly in sinusoids (1.5 ± 0.67; P < 0.05) and in perivenular hepatocytes (0.25 ± 0.86; P < 0.05). Additionally, we also observed a decreased ICAM‐1 reactivity in portal hepatocytes (0.25 ± 0.62), but these differences did not reach statistical significance. Remarkably, after treatment, hepatocytes did not show ICAM‐1 reactivity in resolved AR, but in corticoid‐resistant patients AR did not change or increase. In conclusion, in patients with LT and AR, ICAM‐1 was expressed in hepatocytes and with more intensity in sinusoid cells. Additionally, a down‐regulation of the ICAM‐1 tissue expression after corticoid treatment may exist, although in corticoid‐resistant AR no modulation on ICAM‐1 tissue expression was observed.     

Abdominal peritoneum as a defense organ: Analysis of ICAM‐1 expression in the LPS‐stimulated rat

To investigate the immune environment of the peritoneal cavity, ICAM‐1 (intercellular adhesion molecule) expression on the apical surface of the hepatic peritoneum of LPS (lipopolysaccharide) stimulated rats was anlayzed ultrastructurally and chronologically with immnunoTEM&SEM. ICAM‐1 expression was restricted to the side of microvilli of the mesothelial cells. Microvilli demonstrated bulbous tips and included fuzzy coats and strands. Bulbous tips sometimes expressed the antigen, but fuzzy coats and strands did not. Intervillar cell surfaces lacked its expression. Although ICAM‐1 expression increased eightfold 24 hr after stimulation, the selective expression remained unchanged. These results suggest that microvilli are closely associated with cell migration in the peritoneal cavity through adhesion molecules that establish a road for migration. Clin. Anat. 12:20–26, 1999. © 1999 Wiley‐Liss, Inc.     

ICAM‐1 and B7‐1 provide similar but distinct costimulation for CD8+ T cells,while CD4+ T cells are poorly costimulated by ICAM‐1

The function of purified ICAM‐1 in costimulating CD4⁺ and CD8⁺ T cell responses has been directly compared to that of B7‐1 in a model system that minimizes contributions of other receptor‐ligand interactions. While B7‐1 costimulates both subsets of T cells, ICAM‐1 is much more effective in the costimulation of CD8⁺ cells. ICAM‐1 also synergizes with B7‐1 for the induction of IL‐2 production in CD8⁺ but not CD4⁺ T cells. These differences are not explained by differences in LFA‐1 receptor expression on the two subsets of T cells. The CD8⁺ T cell response to ICAM‐1 costimulation is associated with increased proliferation and IL‐2 production at levels similar to those seen with B7‐1 costimulation, but clonal expansion in response to ICAM‐1 is not as great due to decreased cell survival. ICAM‐1‐mediated costimulation is effective for both naive and memory CT8⁺ T cells, is independent of CD28 engagement, and does not appear to be due solely to effects on adhesion. These results suggest that ICAM‐1‐dependent, B7‐independent costimulation may be important in initiating a CTL response to class I antigen presented by cells that are not professional APC.     

The Influence of Sam-Chil-Geun (Panax Notoginseng) on the Serum Lipid Levels and Inflammations of Rats with Hyperlipidemia Induced by Poloxamer-407

Purpose

Atherosclerosis is characterized by the progressive deposition of lipids and inflammatory process. We attempted to develop a chemically-induced hyperlipidemic mice model, using poloxamer-407 and evaluated the lipid lowering and anti-inflammatory effect of P. notoginseng compared with that of atorvastatin, an antihyperlipidemic drug.

Materials and Methods

Male Wistar rats were randomly divided into 5 groups: control group without any intervention (normal), poloxamer 500 mg/kg i.p. (P), poloxamer plus atorvastatin 1.34 mg/kg p.o. (P + ST), poloxamer plus P. notoginseng 40 mg/kg p.o. (P + NG40), and poloxamer plus P. notoginseng 100 mg/kg p.o. (P + NG100). After 3 weeks, we measured serum total cholesterol, low-density lipoprotein (LDL), high-density lipoprotein (HDL), triglyceride, interleukin (IL)-1, tumor necrosis factor (TNF)-α levels, and reports of cyclo-oxygenase (COX)-2 & intercellular adhesion molecule (ICAM) appearances in each group.

Results

After 3 weeks, serum cholesterol levels significantly decreased in P + ST and P + NG40 groups. Significant decrease of LDL level was only noted in the P + ST group. P + ST, P + NG40, and P + NG100 also had decreased serum triglyceride levels; however, P + ST and P + NG40 showed no statistical difference of the triglyceride lowering effect. The results of IL-1 and TNF-α and the appearance of COX-2 and ICAM were statistically not different in each group.

Conclusions

P. notoginseng 40 mg/kg showed significantly lowering effects on serum total cholesterol and triglyceride levels. We suggest a well-designed study showing the effects of regulating blood lipids with combined administration of P. notoginseng and statin-drug.     

United airways: circulating Th2 effector cells in an allergic rhinitis model are responsible for promoting lower airways inflammation

Background Allergic rhinitis (AR) and asthma often coexist and are referred to as ‘united airways’ disease. However, the molecular and cellular pathways that are crucially involved in the interaction between upper and lower airways remain to be identified. Objective We sought to assess whether and how AR exacerbates lower airway inflammation upon allergen challenge in mice. Methods We previously developed an intranasal ovalbumin (OVA)‐driven AR model, characterized by nasal eosinophilic inflammation, enhanced serum levels of OVA‐specific IgE and Th2 cytokine production in cervical lymph nodes. In OVA‐sensitized mice with or without AR, a lower airway challenge was given, and after 24 h, lower airway inflammation was analysed. Results We found that AR mice were more susceptible to eosinophilic inflammation following a lower airway OVA challenge than OVA‐sensitized controls. AR mice manifested increased numbers of eosinophils in bronchoalveolar lavage fluid and increased inter‐cellular adhesion molecule‐1 (ICAM‐1) expression on lung endothelium, when compared with OVA‐sensitized controls. Depletion of T cells in OVA‐challenged AR mice completely abrogated all hallmarks of lower airway inflammation, including enhanced IL‐5 and tissue eosinophilia. Conversely, adoptive transfer of Th2 effector cells in naïve animals induced lower airway eosinophilic inflammation after challenge with OVA. Blocking T cell recirculation during AR development by the spingosine‐1 analogue FTY720 also prevented lower airway inflammation including ICAM‐1 expression in AR mice upon a single lower airway challenge. Conclusion Our mouse model of ‘united airways’ disease supports epidemiological and clinical data that AR has a significant impact on lower airway inflammation. Circulating Th2 effector cells are responsible for lung priming in AR mice, most likely through up‐regulation of ICAM‐1. Cite this as: A. KleinJan, M. Willart, M. van Nimwegen, K. Leman, H. C. Hoogsteden, R.W. Hendriks and B.N. Lambrecht, Clinical & Experimental Allergy, 2010 (40) 494–504.     

Migration of immature mouse DC across resting endothelium is mediated by ICAM-2 but independent of beta2-integrins and murine DC-SIGN homologues

Immature dendritic cells (DC) reside in tissues where they initiate immune responses by taking up foreign antigens. Since DC have a limited tissue half-life, the DC pool in tissues has to be replenished constantly. This implies that precursor/immature DC must be able to cross non-activated endothelium using as yet unknown mechanisms. Here we show that immature, but not mature bone marrow-derived murine DC migrate across resting endothelial monolayers in vitro. We find that endothelial intercellular adhesion molecule-2 (ICAM-2) is a major player in transendothelial migration (TEM) of immature DC, accounting for at least 41% of TEM. Surprisingly, the ICAM-2-mediated TEM was independent of beta2-integrins, the known ICAM-2 ligands, since neither blocking of beta2-integrins with antibodies nor the use of CD18-deficient DC affected the ICAM-2-specific TEM. In humans, the C-type lectin DC-specific ICAM-3-grabbing nonintegrin (DC-SIGN) was shown to interact with ICAM-2, suggesting a similar role in mice. However, we find that none of the murine DC-SIGN homologues mDC-SIGN, murine DC-SIGN-related molecule-1 (mSIGN-R1) and mSIGN-R3 is expressed on the surface of bone marrow-derived mouse DC. Taken together, this study shows that ICAM-2 strongly supports transmigration of immature DC across resting endothelium by interacting with ligands that are distinct from beta2-integrins and DC-SIGN homologues.     

Inactivated Sendai virus particle upregulates cancer cell expression of intercellular adhesion molecule‐1 and enhances natural killer cell sensitivity on cancer cells

We have already reported that the inactivated Sendai virus (hemagglutinating virus of Japan; HVJ) envelope (HVJ‐E) has multiple anticancer effects, including induction of cancer‐selective cell death and activation of anticancer immunity. The HVJ‐E stimulates dendritic cells to produce cytokines and chemokines such as β‐interferon, interleukin‐6, chemokine (C‐C motif) ligand 5, and chemokine (C‐X‐C motif) ligand 10, which activate both CD8⁺ T cells and natural killer (NK) cells and recruit them to the tumor microenvironment. However, the effect of HVJ‐E on modulating the sensitivity of cancer cells to immune cell attack has yet to be investigated. In this study, we found that HVJ‐E induced the production of intercellular adhesion molecule‐1 (ICAM‐1, CD54), a ligand of lymphocyte function‐associated antigen 1, in several cancer cell lines through the activation of nuclear factor‐κB downstream of retinoic acid‐inducible gene I and the mitochondrial antiviral signaling pathway. The upregulation of ICAM‐1 on the surface of cancer cells increased the sensitivity of cancer cells to NK cells. Knocking out expression of ICAM‐1 in MDA‐MB‐231 cells using the CRISPR/Cas9 method significantly reduced the killing effect of NK cells on ICAM‐1‐depleted MDA‐MB‐231 cells. In addition, HVJ‐E suppressed tumor growth in MDA‐MB‐231 tumor‐bearing SCID mice, and the HVJ‐E antitumor effect was impaired when NK cells were depleted by treatment with the anti‐asialo GM1 antibody. Our findings suggest that HVJ‐E enhances NK cell sensitivity against cancer cells by increasing ICAM‐1 expression on the cancer cell surface.     

怀牛膝总皂苷对大鼠血管平滑肌细胞内bFGF和ICAM-1表达的影响

目的：探讨怀牛膝总皂苷（ABS）对大鼠血管平滑肌细胞（VSMC）内细胞因子的影响,阐明其抗动脉粥样硬化的机制。方法：采用氧化型低密度脂蛋白（OX-LDL）诱导平滑肌细胞增殖的方法,以GAPDH作内标,RT-PCR检测相关细胞因子的表达量,评价ABS对细胞内碱性成纤维细胞生长因子（bFGF）和细胞间黏附分子（ICAM-1）mRNA表达量的影响。结果：OX-LDL浓度为30mg／L促进VSMC增殖最强;ABS（50、100、150mg／L）显著抑制OX-LDL诱导VsMc内bFGF、ICAM-1基因表达,作用呈浓度依赖性。结论：ABS能抑制在OX-LDL作用下VSMC内细胞因子的表达,说明其具有抗动脉粥样硬化的作用。