Hepatic ischemia/reperfusion injury in P-selectin and intercellular adhesion molecule-1 double-mutant mice

Neutrophil adhesion and recruitment represents one of the early cellular events that occur during hepatic ischemia/reperfusion (IR) injury and plays a critical role in determining the extent of tissue damage. The adhesion molecules, such as selectins and intercellular adhesion molecules (ICAM), are important in mediating neutrophil-endothelial cell interactions and neutrophil emigration. The goal of this study was to evaluate the role of P-selectin and ICAM-1 in hepatic IR injury. Male wild-type and P-selectin/ICAM-1-deficient (P/I null) mice underwent 90 minutes of partial hepatic ischemia followed by reperfusion at various time points (0, 1.5, 3, and 6 hours). Reperfusion caused a time-dependent hepatocellular injury in both wild-type and P/I null mice as judged by plasma alanine aminotransferase (ALT) levels and liver histopathology examination. Although ALT levels were slightly lower in the P/I null mice compared with the wild-type mice the differences were not statistically significant. Neutrophil infiltration to the ischemic liver was observed in both mouse groups after 6 hours of reperfusion; however, the infiltration to the midzonal region of the ischemic liver was more pronounced in the wild-type group. This study suggests that hepatocellular injury induced after hepatic IR was independent of P-selectin and ICAM-1 in this model of acute inflammatory tissue injury.     

脊髓缺血-再灌注损伤细胞间粘附分子-1及白细胞介素-1β的表达

目的:探讨脊髓缺血-再灌注损伤细胞间粘附分子-1(ICAM-1)及白细胞介素-1β(IL-1)的表达对血脊髓屏障损害的分子机制。方法:将77只同龄Wistar大鼠,随机分为正常对照组、单纯缺血组和缺血再灌组。手术方法采用Zivin法复制模型。应用逆转录-聚合酶链反应、地高辛标记cDNA探针技术、免疫组化及免疫荧光激光共聚焦扫描显微镜技术检测脊髓再灌注损伤血管内皮ICAM-1mRNA和IL-1βmRNA表达量。结果:正常组和单纯缺血组未引起细胞因子和粘附分子表达量的增加。而缺血再灌注后缺血区细胞因子、粘附分子的表达及多形核白细胞(PMN)的浸润先后发生了改变。再灌注2h,IL-1βmRNA的表达首先升高,约为对照组的2倍。再灌注6h达到高峰,并持续到12h。ICAM-1mRNA表达量于再灌注4h明显升高,再灌注12h其在单位微血管面积上的荧光强度约比单纯缺血组增加了1/2。结论:再灌注损伤后脊髓微血管内皮ICAM-1及其调节因子IL-1β的表达量增加是导致血脊髓屏障损害的重要分子基础。     

细胞间粘附分子-1的反义寡核苷酸在小鼠心脏移植中的作用

目的 探讨细胞间粘附分子-1(ICAM-1)的反义寡核苷酸(ICAM-1-ASO)在小鼠心脏移植中对ICAM-1表达以及在排斥反应中的作用.方法 以BALB/c小鼠为供者,C57小鼠为受者,建立颈部异位心脏移植模型,移植后将受者随机分为3组,每组10只.分别为单纯移植组、对照寡核苷酸组(对照ODN)和ICAM-1-ASO组.各组移植后24 h开始经受者静脉分别注射双蒸水0.3 ml、对照ODN 10 mg/kg和ICAM-1-ASO 10 mg/kg,每天1次,连续5 d.移植后第8天各组随机取5只受者切取移植心,进行病理学检查;采用免疫组织化学方法对移植心组织中ICAM-1的表达进行观察;采用半定量逆转录聚合酶链反应(RT-PCR)对ICAM-1mRNA表达进行定量分析.每组中余下的小鼠继续观察直至移植心停跳,记录各组移植心的存活时间.结果 单纯移植组心脏移植后第8天可观察到明显的排斥反应,移植心心跳减慢、心律不齐、心律紊乱、搏动无力;病理学检查可见大量的淋巴细胞浸润,心肌间质水肿及大片心肌坏死;与移植前对比,内皮细胞和心肌细胞ICAM-1表达明显增强;ICAM-1mRNA表达也明显增加;移植心存活时间平均为(9.8±1.48)d.对照ODN组与单纯移植组差异无统计学意义.而ICAM-1-ASO组与单纯移植组相比,移植心ICAM-1的表达明显减弱,淋巴细胞浸润明显减少,排斥反应程度减轻;移植后第9天心律基本规整,偶有心律不齐,心跳力度较单纯移植组增强;移植心存活时间明显延长至(14.6±1.14)d.结论 ICAM-1-ASO抑制移植心组织中ICAM-1的表达和淋巴细胞的浸润,抑制移植后排斥反应的发生和发展,改善移植物的功能,延长移植物的存活时间,其作用具有序列特异性.     

细胞间粘附分子-1的反义寡核苷酸在异种心脏移植中的作用

目的探讨细胞间粘附分子1的反义寡核苷酸(ICAM1ASO)在小鼠→大鼠的异种心脏移植中对ICAM1表达以及排斥反应的作用。方法BALB/c小鼠和Lewis大鼠分别作为供者和受者,随机配对建立小鼠→大鼠的异种心脏移植模型。按照供者术前处理不同分为双蒸馏水对照组、寡核苷酸对照组和ICAM1ASO组。各组供者术前6h分别从右颈外静脉注射双蒸馏水、寡核苷酸和ICAM1ASO。术后48h各组取5只受者切取供心进行病理学检查;采用免疫组织化学方法对供心组织ICAM1的表达进行观察;半定量逆转录聚合酶链(RTPCR)法对ICAM1mRNA的表达进行定量分析。每组中余下5只受者继续观察直至供心停跳,记录供心存活时间。结果双蒸馏水对照组和寡核苷酸对照组在移植后48h可观察到供心心跳减慢,出现不同程度的心律不齐、心律紊乱,供心搏动无力;供心病理学检查显示心脏组织中广泛间质充血、水肿、出血及血栓形成,部分心肌髓样变性及溶解并伴有炎性细胞浸润;与移植前BALB/c小鼠心脏对比,内皮细胞和心肌细胞ICAM1表达明显增强;ICAM1mRNA表达也明显增加;供心存活时间分别为(53.6±3.58)h和(55.0±3.16)h。ICAM1ASO组供心ICAM1在蛋白水平和mRNA水平的表达均减弱;排斥反应程度也明显减轻;供心存活时间为(66.4±2.61)h;与寡核苷酸对照组和双蒸馏水对照组比较,差异有统计学意义。结论ICAM1ASO能明显抑制异种移植中供心ICAM1的表达,抑制异种移植后排斥反应的发生和发展,延长移植物的存活时间,其作用具有高度序列特异性。     

P选择素、细胞间黏附分子-1在大鼠胰腺移植缺血再灌注损伤中的作用

目的观察P选择素（Ps）和细胞间黏附分子-1（ICAM-1）在大鼠胰腺移植缺血再灌注损伤中的作用及Ps单克隆抗体的治疗作用。方法75只SD大鼠随机分为假手术组、移植组和治疗组。移植组和治疗组均进行全胰十二指肠移植术，但治疗组于再灌注前5rain静脉注射Ps单克隆抗体。3组分别于腹主动脉开放后1（n=5）、3（n=5）、6（n=5）h取血测定血清淀粉酶水平，并切取胰腺标本进行组织病理学观察及Ps、ICAM．1免疫组织化学染色。结果移植组胰腺组织损伤随再灌注时间的延长而加重，血淀粉酶升高，与中性粒细胞浸润直接相关；而治疗组胰腺组织损伤不明显，血淀粉酶减低。移植组各时段Ps、ICAM-1均有阳性表达，且Ps再灌注1h为表达高峰，ICAM．1随再灌注时间延长表达逐渐增加，假手术组、治疗组Ps、ICAM-1不表达。结论Ps及ICAM-1按一定时间顺序参与胰腺移植缺血再灌注损伤的病理过程；Ps可能是胰腺移植缺血再灌注损伤的起始因素；抗Ps单克隆抗体对移植胰腺缺血再灌注损伤有保护作用。     

Methoxyethyl-modified intercellular adhesion molecule-1 antisense phosphorothiateoligonucleotides inhibit allograft rejection, ischemic-reperfusion injury, and cyclosporine-induced nephrotoxicity

BACKGROUND: The addition of phosphorothioate (PS) groups to natural phosphodiester (PD) antisense oligodeoxynucleotides (oligo) prevents their in vivo hydrolysis by nucleases allowing an RNase-dependent elimination of targeted mRNA. To further improve oligo function 2'-methoxyethyl (ME) groups were attached to selected nucleotides at the 3'-end because ME groups block RNase activity. METHODS/RESULTS: ME modification of PS- or PD/PS-oligo targeting human intracellular adhesion molecule (ICAM)-1 mRNA significantly increased the degree and duration of the in vitro inhibitory effects without compromising selectivity and specificity. A 7-day intravenous or oral therapy with rat ME/PS-modified ICAM-1 antisense oligo extended the survivals of kidney allografts. In addition, ME/PS-modified ICAM-1 antisense oligo reduced ischemic-reperfusion injury in kidneys, as measured by glomerular filtration rate, creatinine levels, and infiltration with leukocytes. Finally, a 14-day treatment with cyclosporine (CsA)-induced nephrotoxicity in syngeneic kidney transplants correlated with both increased ICAM-1 protein expression and infiltration with leukocytes. Graft perfusion and treatment of recipients with ICAM-1 antisense ME/PS-oligo alleviated the nephrotoxic effect and decreased ICAM-1 expression and leukocyte infiltration. CONCLUSIONS: ME/PS-modified ICAM-1 antisense oligo is very effective in inhibiting the ICAM-1-dependent mechanism of graft infiltration and tissue damage involved in allograft rejection, ischemic-reperfusion injury, and CsA-induced nephrotoxicity.     

Intercellular adhesion molecule-1 expression in human kidneys with glomerulonephritis.

Intercellular adhesion molecule-1 (ICAM-1) plays an important role in immune responses. Expression of this molecule was examined on cryostat sections of 15 normal human and 112 glomerulonephritic kidneys using a specific monoclonal antibody and the indirect immunoperoxidase staining technique. The expression of ICAM-1 on renal structures was compared to that of HLA-DQ, -DR, -DR/DP and -DP antigens. On normal kidneys ICAM-1 was observed on some Bowman's capsular cells and on single glomerular cells which probably represent endothelial and mesangial cells. ICAM-1 was present on peritubular capillaries and on vascular endothelium of large vessels as well as on fibroblasts, whereas no expression of ICAM-1 on proximal tubular epithelial cells (PTECs) was detected. In normal renal tissues the distribution of ICAM-1 was similar to that of HLA-DQ and -DP antigens. In kidneys with different forms of glomerulonephritis especially in association with interstitial inflammation, an abnormal expression of ICAM-1 on PTECs was frequently correlated with aberrant expression of HLA-DQ and -DP antigens. These results further support the hypothesis that PTECs may participate in cell-mediated immune reactions in glomerulonephritis.     

组织细胞间粘附分子-1在阻塞性黄疸小肠粘膜损伤中的作用及丹参防治机制的研究

目的：研究组织细胞间粘附分子－1（ICAM－1）在阻塞性黄疸（阻黄）小肠粘膜损伤中的作用及丹参防治小肠粘膜损伤机制。方法：SD大鼠48只分为4组：假手术对照组（SO＋NS）、阻黄组（BDL＋NS）、治疗对照组（SO＋SM）及丹参治疗组（BDL＋SM），每组术后再随机分设7、14d两个时相点。在不同时相点检测小肠组织髓过氧化物酶活性（MPO）、ICAM－1、二胺氧化酶（DAO）、血浆内毒素水平变化，并与丹参治疗组进行比较。结果：BDL＋NS组7、14d时小肠DAO的活性降低，MPO活性增高P＜0．05），门表脉血浆内毒素增加，ICAM－1主要表达在小肠粘膜上皮组织，且表达逐渐增强（P＜0．05）；BDL＋SM组7、14d时小肠组织DAO活性显著升高，门静脉血浆内毒素降低，ICAM－1表达受到抑制（P＜0．05），MPO改变不明显。结论：阻黄引起小肠上皮细胞上的ICAM－1表达上调，参与了中性粒细胞（PMN）介导的小肠粘膜损伤；丹参是通过抑制ICAM－1的表达而减轻小肠粘膜的损伤。     

Attenuation of cytomegalovirus-induced endothelial intercellular adhesion molecule-1 mRNA/protein expression and T lymphocyte adhesion by a 2'-O-methoxyethyl antisense oligonucleotide

BACKGROUND: Intercellular adhesion molecule-1 (ICAM-1) is strongly induced under inflammatory conditions associated with allograft rejection, thereby promoting leukocyte recruitment and activation at the site of inflammation. Enhancement of ICAM-1 expression can also be the result of viral infection, in particular human cytomegalovirus (CMV), a frequent source of complications in the transplant recipient. In vitro studies have shown that CMV infection of endothelial cells (EC) results in the direct enhancement of ICAM-1 expression and consequent leukocyte adhesion/activation suggesting mechanisms by which CMV exacerbates graft vascular disease. Although treatment of EC with ICAM-1-specific antisense oligonucleotides has been shown to attenuate ICAM-1 induction under simulated inflammatory conditions (i.e., TNF-alpha), no studies have addressed their effectiveness on virally-induced ICAM-1 expression. RESULTS: In the current investigation, we show that the progressive increase in endothelial ICAM-1 protein expression that follows inoculation with CMV correlates with a progressive accumulation of ICAM-1 mRNA. Furthermore, we demonstrate that treatment of EC with a partially 2'-O-methoxyethyl modified ICAM-1-specific antisense oligonucleotide before viral inoculation significantly reduces CMV-associated induction of ICAM-1 protein and mRNA expression. Finally, we show that antisense-mediated attenuation in ICAM-1 expression results in a significant reduction of T lymphocyte adhesion to CMV-infected EC monolayers, an interaction that has been implicated in allogeneic T lymphocyte activation, in viral transmission to transiently adherent leukocytes and subsequent hematogenous dissemination. CONCLUSIONS: These findings demonstrate for the first time that antisense oligonucleotides can effectively reverse virally-induced host cellular protein expression, specifically ICAM-1, as well as consequent T lymphocytes adhesion, thus broadening the potential clinical utility of antisense oligonucleotides.     

Mesangial expression of intercellular adhesion molecule-1 in primary glomerulosclerosis.

Frozen sections of renal biopsy specimens from eight patients with primary focal segmental glomerulosclerosis (FSGS) and 10 patients with membranous nephropathy (MN) were stained in immuno-peroxidase with the intercellular adhesion molecule-1 (ICAM-1) monoclonal antibody (MoAb), CL203.4. ICAM-1 was expressed by mesangial cells in six patients with FSGS. On the other hand, ICAM-1 was not detected in mesangial cells in patients with MN or in the non-affected portion of tumoral kidneys used to control normal renal expression of ICAM-1. De novo mesangial expression of ICAM-1 in FSGS suggests that sclerosis results from an inflammatory process, possibly associated with local release of cytokines.     

缺血再灌注对大鼠肺细胞间粘附分子-1表达的影响

目的 观察缺血再灌注（Ｉ／Ｒ）对肺组织细胞间粘附分子－１（ＩＣＡＭ－１）表达的影响，分析ＩＣＡＭ－１表达与肺内中性粒细胞浸润的关系。方法 Ｗｉｓｔａｒ大鼠单肺原位热缺血再灌注损伤模型分７组，左肺缺血９０ｍｉｎ，再灌注时间分别为０、１、２、４、８、１６、２４ｈ。逆转录－聚合酶链式反应（ＲＴ－ＰＣＲ）法及Ｗｅｓｔｅｒｎ ｂｌｏｔ法检测肺组织ＩＣＡＭ－１ｍＲＮＡ及蛋白表达，测定各组肺组织髓过氧化物酶活性     

丙酮酸乙酯对脓毒症肺损伤大鼠细胞间粘附分子-1的影响

目的研究丙酮酸乙酯对脓毒症肺损伤大鼠细胞间粘附分子1(ICAM1)的影响,探讨丙酮酸乙酯保护大鼠脓毒症肺损伤的作用机制。方法采用盲肠结扎穿孔术复制脓毒症肺损伤动物模型,30只Wistar大鼠随机分为假手术组、脓毒症肺损伤组和丙酮酸乙酯(40mg/kg,腹腔内注射)治疗组,每组10只。所有大鼠12h后活杀,用免疫组织化学方法检测肺组织中ICAM1的分布和表达,用免疫印迹法检测肺组织ICAM1蛋白水平的表达,检测支气管肺泡灌洗液白细胞计数、肺湿/干重比和肺组织髓过氧化物酶(MPOase)活性。结果与脓毒症肺损伤组比较,丙酮酸乙酯治疗组免疫印迹法和免疫组织化学法显示肺组织ICAM1的表达显著降低,肺泡灌洗液白细胞计数、肺湿/干重比和MPOase活性降低(P<0.01)。结论丙酮酸乙酯通过抑制肺组织ICAM1的表达,对脓毒症肺损伤大鼠具有保护作用。     

Increased expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 in rat cardiac allografts

This study was designed to investigate the role of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) during chronic cardiac allograft rejection. Wistar rats were used as donors, and SD rats as recipients heterotopic cardiac transplants. Recipients pretreated with inoculation of donor splenocytes (SPC) followed by cyclophosphamide (CP) were divided into 4 groups: (A) untreated group (n = 18) without immunosuppression; (B) SPC plus CP-treated group (n = 18) that were euthanized at 15-120 days posttransplantation; (C) CsA-treated group (n = 18) euthanized at 2-3 months posttransplantation; and (D) tolerance group (n = 18) treated with SPC plus CP and monitored for at least 1 year posttransplantation. Cardiac allografts were harvested at various times for immunohistochemical studies performed to evaluate the expression of ICAM-1 and VCAM-1. Pretreatment of animals with SPC and CP induced long-term cardiac allograft survival. Immunohistochemical staining demonstrated a low level of ICAM-1 and VCAM-1 expression in cardiac allograft muscle and coronary arteries among Groups B and D. In contrast, the expressions of ICAM-1 and VCAM-1 in cardiac allografts of Groups A and C were significantly higher than those in Groups B and D. Our results suggested that the expression of ICAM-1 and VCAM-1 plays an important role during the development of chronic cardiac allograft rejection.     

颅脑损伤后血清中sICAM-1和IL-1β含量变化的意义

目的 动态观测急性颅脑损伤患者血清中sICAM-1和IL-1β含量的变化．探讨其临床意义。方法 按入院先后随机选取25例被CT证实的颅脑损伤患者，按照入院时、24．48．96小时四个时间段分别采集血标本，应用双抗体夹心酶联免疫吸附法（ELISA）测定sICAM-1和IL-1β的含量．并与对照组比较。结果 颅脑损伤患者早期sICAM-1含量较对照组无变化，在48小时明显升高，并持续升高至96小时（P〈0．05）。sICAM-1含量与损伤程度无关，而与预后相关。皿清中IL-1β含量于入院时就有明显升高（P〈0．05），IL-1β升高不仅与损伤程度有关，而且与预后也密切相关。血清中IL-1β含量与sICAM-1含量相关。结论 血清中sICAM-1和IL-1β参与了颅脑后继发性脑损伤，sICAM-1和IL-1β升高反映了颅内的炎症反应。