烧伤患者血清对肺毛细血管内皮细胞与中性粒细胞粘附特性的研究

目的　探讨烧伤继发急性肺损伤 (acutelunginjury ,ALI)患者肺毛细血管内皮细胞(PVEC)与中性粒细胞 (PMN)的粘附能力 ,观察C5a在PVEC PMN粘附行为中的作用程度及规律。　方法　采用微管吸吮技术 ,定量测定PVEC PMN之间粘附力值变化 ;测定髓过氧化酶 (MPO)活性 ,反映PVEC粘附PMN的数量。　结果　随着rh C5a浓度上升 ,PVEC粘附了更多的PMN ;粘附力值也逐渐增加 ,3 0 0 μg/Lrh C5a时差异有非常显著性意义 (P <0 .0 1) ;MPO也随之上升 ;工作浓度为 1∶10 4 的anti C5aRMcAb可缓解急性肺损伤 (ALI)血清所致的MPO增加 ,呈量效关系 ,　结论　C5a、C5aR共同参与了PVEC PMN的粘附过程。而且急性肺损伤所致的粘附力值上升可被anti C5aRMcAb所抑制 ,提示C5a、C5aR在烧伤继发的急性肺损伤的发病机制中可能起到了重要作用     

抑制核因子-κB的活性减少内皮细胞与T淋巴细胞粘附

目的探讨抑制核因子κB(NFκB)对内皮细胞活化及内皮细胞与T淋巴细胞粘附的影响。方法应用pCMVIκBαM质粒建立稳定表达突变型IκBα蛋白的内皮细胞系,同时以表达野生型IκBα蛋白(pCMVIκBα)的内皮细胞作为对照。通过逆转录聚合酶链反应及流式细胞仪检测细胞表面细胞间粘附分子1(ICAM1)、血管细胞粘附分子1(VCAM1)和P选择素的变化,并将两种转染细胞分别与人T淋巴细胞株Jurkat细胞混合培养,通过相差显微镜下细胞计数来观察内皮细胞与T淋巴细胞的粘附情况。结果高表达突变型IκBα蛋白的内皮细胞的NFκB活性被显著抑制,其ICAM1mRNA和VCAM1mRNA水平明显低于对照细胞(P<0.05),细胞表面的ICAM1、VCAM1和P选择素的表达较对照细胞显著减少(P<0.05),与T淋巴细胞的粘附较对照细胞显著减少(P<0.05)。结论抑制核因子κB的活性能有效减少内皮细胞与T淋巴细胞的粘附。     

细胞间粘附因子-1在门静脉高压患者脾静脉的表达及意义

目的 探讨细胞间粘附因子—1(ICAM—1)在门静脉高压患者脾静脉的表达及其在贲门周围血管离断术后门静脉血栓形成的意义。方法 对34例门静脉高压患者和34例单纯脾破裂患者的脾静脉行苏木素—伊红染色观察形态学变化；行ICAM—1原位杂交并行定量分析，术后观察门静脉血栓的发生，对两者的关系进行研究。结果 门静脉高压患者脾静脉中膜平滑肌增生，内膜增厚，门静脉高压组和脾破裂组之间脾静脉内皮细胞ICAM—1mRNA表达差异有非常显著性(P＜0．01)；脾破裂组术后无门静脉血栓形成，门静脉高压组有8例门静脉血栓形成；门静脉高压组内有无门静脉血栓形成病例之间脾静脉内皮细胞ICAM—1mRNA表达差异有显著性(P＜0．05)。结论 门静脉高压性血管病变及其内皮细胞ICAM—1mRNA过度表达可能是门静脉高压患者门静脉血栓形成重要原因。     

血小板内皮细胞粘附分子-1在机械通气致肺损伤中的作用

目的：探讨血小板内皮细胞粘附分子－1（PECAM－1）在机械通气致肺损伤中的作用。方法：24只普通健康小猪，随机等分为对照组，低潮气量（A组），正常潮气量组（B组）及大潮气量组（C组），持续给予不同潮气量通气，利用免疫组织细胞化学技术，髓过氧化物酶（MPO）测定法及病理组织切片技术，分别检测不同潮气量组通气1d,3d,7d后肺血管内皮细胞表面PECAM－1表达量，血清和肺组织匀浆中MPO活性的变化及肺血管内皮细胞结构及连接的改变。结果：A，B，C组血清，肺组织匀浆MPO活笥较对照组升高（P＜0．05或0．01），A，B，C组肺血管内皮组织表面PECAM－1对照组表达下调（P＜0．05或0．01），内皮细胞及基膜肿胀，内皮细胞间间隙增大，均以7d后明显，尤以A，C组显著。结论：PECAM－1在机械通气致肺损伤中可能发挥重要作用。     

烟雾吸入伤血清对中性粒细胞粘附及迁移作用的体外研究

目的通过鼠烟雾吸入伤血清体外刺激培养内皮细胞和中性粒细胞(PMN),了解PMN粘附与迁移变化,以及抗CD11a、抗CD11b和抗ICAM-1对PMN粘附与迁移的影响,探讨粘附因子在烟雾吸入性损伤中的作用。方法在体外实验中用荧光标记PMN,测定PMN与烟雾吸入性损伤血清刺激的内皮细胞的粘附率,在5μm滤膜上培养内皮细胞测定PMN迁移量,并采用抗体阻断技术,测定了粘附因子CD11a、CD11b和ICAM-1在PMN粘附和迁移中的作用。结果内皮细胞在此血清中培养12～24小时,PMN粘附率是正常时的3倍多,在此血清中培养24小时时PMN迁移量也增至2倍多,表明吸入伤血清能促进PMN与内皮细胞粘附和PMN通过毛细血管内皮向血管外迁移;PMN迁移量与粘附率呈显著正相关(r=0.975,P<0.01)。PMN和内皮细胞粘附因子抗体(抗CD11a、抗CD11b和抗ICAM-1)能减少PMN与内皮细胞粘附率(44％、55％和51％)和PMN迁移量(39％、65％和58％)。结论烟雾吸入伤血清能增加PMN的粘附和迁移,其可能是PMN浸润的病理基础。进一步研究证明,粘附因子CD11a、CD11b和ICAM-1的增加在PMN浸润过程中起重要作用,而且浸润的PMN释放氧自由基和蛋白酶能导致进一步的肺组织损害。     

己酮可可碱对淡水淹溺后肺粘附分子表达的影响

目的　探讨己酮可可碱 (pentoxifylline ,PTX)对淡水淹溺肺损伤后肺组织细胞间粘附分子 (ICAM 1 )、血管间粘附分子 (VCAM 1 )表达的影响。方法　 1 8只普通杂种犬随机分为对照组、淹溺组和淹溺加己酮可可碱组三组。采用气管切开插入Carlen’s管分隔左右两肺 ,向右肺灌淡水2 0ml/kg ,左肺自主通气的方法模拟淡水淹溺造成直接和间接肺损伤模型。于淹溺后 1、2、4、6、8h五个观察时间点采集肺组织 ,逆转录 聚合酶链 (RT PCR)方法检测肺组织中粘附分子的表达 ,光镜下病检测中性粒细胞 (polymorphonuclearneutrophil,PMN)在肺组织的浸润情况。 结果　淹溺后各时相点肺组织中性粒细胞大量浸润 ,ICAM 1、VCAM 1mRNA表达均有显著的升高。ICAM 1mRNA淹溺后 4h达高峰为 (87 9± 6 1 ) % (P <0 0 1 ) ,6h后开始下降。PTX则可以明显抑制这种变化 (P<0 0 1 )。同样VCAM 1mRNA有微量表达 ,PTX组比淹溺组相应时相点表达减少 (P <0 0 1 )。结论　PTX可以通过降低肺淹溺后的ICAM 1、VCAM 1的表达 ,从而减轻PMN在肺内的聚集、活化 ,防止淡水淹溺后急性肺损伤     

粘附分子基因表达在大鼠急性坏死性胰腺炎肺损伤中的作用

目的　研究急性坏死性胰腺炎 (ANP)大鼠肺组织细胞间粘附分子 1(ICAM 1)mRNA的表达情况及其与肺损伤的关系。方法　将 33只Wistar大鼠随机分为正常对照组和胰腺炎不同时间点 (1、4、12和 2 4h)各组 ,用逆行胰胆管注射方法制备ANP模型。采用逆转录聚合酶链反应法检测ANP大鼠肺组织中ICAM 1mRNA表达 ,同时观察肺组织髓过氧化物酶 (MPO)及病理改变。结果造模ANP 1h后大鼠肺组织中ICAM 1mRNA(0 82± 0 0 3)比正常对照组 (0 41± 0 0 4)的表达水平高 (P <0 0 5 ) ,并持续升高至 12h及 2 4h(分别为 1 17± 0 0 5及 1 11± 0 0 4) ,同时伴有肺组织病理损害 ,其严重程度与ICAM 1mRNA表达、MPO及肺MPO与ICAM 1mRNA表达均呈正相关 ,相关系数分别为0 85、0 85及 0 96 (P <0 0 5 )。结论　ANP大鼠早期肺组织中ICAM 1mRNA呈过度表达 ,肺损伤严重程度与ICAM 1mRNA表达的高低有关。肺组织中ICAM 1过度表达及中性粒细胞浸润是ANP肺损害发生的原因之一。     

中性粒细胞在ARDS中的作用

ARDS基本的病理生理变化是自身免疫系统介导的肺泡毛细血管膜损伤。细胞动力学研究表明,血管中50～60%中性粒细胞储存在肺血管床,存在“着边”和沿血管内膜“滚动”现象。由于肺毛细血管的物理约束力,其绝大部分滞留在肺毛细血管。当失血性或感染性休克时,肺血流成慢,肺血管容量进一步扩大。随之,炎性调节因子介导的粘附机制增加中性粒细胞聚集。参与内毒素反应的细胞因子主要有TNF-α、Il-1、Il-8和Il-6。炎性内皮细胞增加其表面内皮细胞白细胞粘附分子(ELAM),细胞间粘附分子(ICAM),以及细胞颗粒膜蛋白140(GTMP-140)。未受刺激的中性粒细胞表面存在植物凝血素粘附分子(LECAM),活化的中性粒细胞则表达整合素(Intergrins)。整合素是由普通β_2单位或一组不同的18链(CD18)与不同的α链(CDlla、CDllb、CDllc)共价结合所组成。在其表面形成三个不同的异构二聚体。一般中性粒细胞粘附和     

p38 MAPK在LPS诱导内皮细胞表达ICAM-1中的作用

目的 研究p38丝裂原激活蛋白激酶(MAPK)信号转导通路在脂多糖(LPS)诱导人脐静脉内皮细胞(HUVEC)表达细胞间粘附分子-1(ICAM—1)中的作用。方法 脐静脉内皮细胞培养后分为两组：(1)刺激组,设不同时相点分别用LPS刺激内皮细胞;(2)预处理组,在LPS刺激前2h,用SB203580预处理内皮细胞。观察ICAM—1蛋白和mRNA表达的变化,检测内皮细胞p38MAPK活性变化。结果 LPS刺激后,内皮细胞表面ICAM—1分子在8～36h显增加,胞浆中mRNA在2h即有显增加;LPS刺激HUVEC后15min,p38MAPK活性即有升高,30～60min达高峰。p38抑制剂SB203580可显抑制LPS的诱导作用。结论 LPS可能通过激活p38MAPK信号转导通路,调节HUVEC的ICAM—1基因和蛋白表达。     

豚鼠哮喘模型单个核细胞及嗜酸细胞与肺微血管内皮细胞粘附性的研究

目的观察哮喘豚鼠模型嗜酸细胞和淋巴细胞与肺微血管内皮细胞的粘附率,探讨肺微血管内皮细胞在哮喘模型发病中的功能变化及炎症细胞与其粘附的分子基础.方法复制豚鼠哮喘模型;分离、培养和鉴定肺微血管内皮细胞;转染核因子(NF)-κB反义寡核苷酸;分别用对照组血清和模型组血清孵育;分离外周血淋巴细胞和嗜酸细胞;采用RT-PCR法测定内皮细胞和血管细胞粘附分子-1(VCAM-1)及嗜酸细胞趋化因子(eotaxin)mRNA的表达强度,EMSA法测定内皮细胞NF-κB、活化蛋白(AP-1)的DNA结合活性.结果(1)肺微血管内皮细胞在模型组血清刺激下,与两组外周血淋巴细胞及仅与模型组嗜酸细胞的粘附率显著升高,内皮细胞与嗜酸细胞两者的激活,在嗜酸细胞粘附中起明显的协同作用;(2)肺微血管内皮细胞经模型组血清刺激后,VCAM-1和eotaxin mRNA表达量显著增多,NF-κB、AP-1的DNA结合活性也显著升高.结论肺微血管内皮细胞可被哮喘模型血清激活,使粘附分子、趋化因子及核转录因子的合成和分泌增多,从而显著提高其与炎症细胞的粘附率,核转录因子NF-κB和AP-1对内皮细胞合成炎症因子可能起调控作用.淋巴细胞与嗜酸细胞的粘附机制不尽相同.     

Role of CO-releasing molecules liberated CO in attenuating leukocytes sequestration and inflammatory responses in the lung of thermally injured mice

BACKGROUND: Acute lung injury and pulmonary inflammatory responses are important complications most frequently encountered in severely burned patients. Polymorphonuclear leukocyte (PMN) sequestration and the subsequent generation of oxidants and inflammatory mediators play the key roles in the pathogenesis of acute lung injury. In this study, we used CO-releasing molecules (CORM-2) to determine whether the CO-releasing molecules-liberated CO could attenuate leukocytes sequestration and the inflammatory response in the lung of thermally injured mice. MATERIALS AND METHODS: Fifty-four mice were assigned to three groups in three respective experiments. In each experiment, mice in sham group (n=6) underwent sham thermal injury, whereas mice in the burn group (n=6) received 15% total body surface area (TBSA) full-thickness thermal injury and mice in CORM-2 group (n=6) underwent the same thermal injury with immediate administration of CORM-2 (8 mg/kg, i.v.). PMN accumulation (MPO assay) in mice lungs and tumor necrosis factor-alpha and interleukin-1beta in BAL fluid, pulmonary edema formation, and wet/dry weight ratios of lung were determined. Activation of NF-kappaB and expression level of ICAM-1 in the lung was assessed. In in vitro experiment, PMN adhesion to experimental mice serum-stimulated mouse lung endothelial cells (MLEC) was assessed. RESULTS: Treatment of thermally injured mice with CORM-2 attenuated PMN accumulation and prevented activation of NF-kappaB in the lung. This was accompanied by a decrease of the expression of ICAM-1. In parallel, PMN adhesion to MLEC stimulated by CORM-2-treated thermally injured mice serum was markedly decreased. Also, CORM-2 markedly decreased the production of inflammatory mediators in BAL fluid without suppressing the permeability of pulmonary microcirculation. CONCLUSIONS: CORM-released CO attenuates the inflammatory response in the lung of thermally injured mice by decreasing leukocyte sequestration and interfering with NF-kappaB activation, protein expression of ICAM-1, and therefore, suppressing endothelial cells' pro-adhesive phenotype.     

外源性一氧化碳释放分子2对严重烧伤小鼠肝脏炎性反应的抑制作用

目的 观察外源性一氧化碳释放分子2（CORM-2）对严重烧伤小鼠肝脏炎性反应的抑制作用，并探讨其机制。方法 将C57BL／6小鼠随机分成假伤组（模拟烧伤）、假伤＋CORM-2组、烧伤组、烧伤＋CORM-2组及烧伤＋二甲亚砜（DMSO）组，每组9只。假伤＋CORM-2组除伤后使用CORM-2以外，其他处理同假伤组。烧伤＋CORM-2组及烧伤＋DMSO组除伤后分别使用CORM-2、DMSO外，其余处理同烧伤组。于伤后24h检测小鼠血清丙氨酸转氨酶（ALT）及天冬氨酸转氨酶（AST）的水平，肝组织髓过氧化物酶（MPO）及核因子KB（NF-kB）活性，胞间黏附分子1（ICAM-1）和血管细胞黏附分子1（VCAM-1）蛋白的表达；检测各组小鼠肝窦内皮细胞（HSEC）经各自血清刺激后对中性粒细胞（PMN）的黏附作用。结果 与假伤组比较，烧伤组小鼠血清ALT、AST的水平[（398±34）、（122±22）U／L]及肝组织MPO活性、肝组织ICAM-1和VCAM-1蛋白表达水平均明显升高（P〈0．05或P〈0．01）。与烧伤组比较，烧伤＋CORM-2组上述情况明显改善，且NF-kB活性下降。与假伤组比较，烧伤组小鼠HSEC对PMN的黏附作用增强；烧伤＋CORM-2组该作用明显弱于烧伤组（P〈0．05）。结论 外源性CORM-2能明显抑制肝组织NF-kB活性，减少ICAM-l、VCAM-l蛋白的表达水平，减轻严重烧伤后组织中白细胞滞留，改善肝功能，可有效减轻肝脏炎性反应。     

烟雾吸入伤血清对中性粒细胞粘附及迁移作用的体外研究

目的 通过鼠烟雾吸入伤血清体外刺激培养内皮细胞和中性粒细胞（ＰＭＮ），了解ＰＭＮ粘会与迁移变化，以及抗ＣＤ１１ａ，抗ＣＤ１１ｂ和抗ＩＣＡＭ－１对ＰＭＮ粘附与迁移的影响，探讨粘附因子在烟雾吸入性损伤中的作用。方法 在体外实验中用荧光标记ＰＭＮ，测定ＰＭＮ与烟雾吸入性损伤血清刺激的内皮细胞的粘附率，在５μｍ滤膜上培养仙皮细胞测定ＰＭＮ迁移量，并采用抗体阻断技术，测定了粘附因子ＣＤ１１ａ，ＣＤ１１ｂ和Ｉ     

严重烧伤早期粘附分子的表达对中性粒细胞在肝脏中聚集的影响

研究严重烧伤后早期中性粒细胞在肝脏中的聚集机制及相关粘附分子表达变化。结论严重烧伤后早期粘附分子Ｐ－选择素和ＩＣＡＭ－１在肝窦内皮细胞和肝细胞表面的增加是中性粒细胞在肝脏中聚集的关键，介导了中性料细胞对脏功能的损害。     

盐酸戊乙奎醚对大鼠胸部撞击致急性肺损伤及肺组织Toll样受体4表达的影响

目的 探讨盐酸戊乙奎醚对大鼠胸部撞击致急性肺损伤及肺组织Toll样受体4(TLR4)表达的影响.方法 健康雄性SD大鼠96只,体重250～300 g,采用随机数字表法,将大鼠随机分为3组(n=32):对照组(C组)只麻醉,不制备模型;肺损伤组(ALI组);盐酸戊乙奎醚组(PHcD组)模型制备后即刻,腹腔注射盐酸戊乙奎醚2 mg/kg.砝码(300g)于95 cm高处自由落体撞击大鼠心前区以制备急性肺损伤模型.于模型制备后2、8、12和24h时取8只大鼠,取动脉血样,测定血清TNF-α浓度.于模型制备后8 h取8只大鼠,取动脉血样,行动脉血气分析,随后处死大鼠,取肺组织观察病理学结果,测定干/湿重比(W/D比)、髓过氧化物酶(MPO)活性和TLR4表达水平.结果 与c组比较,ALI组和PHCD组pH值和PaO2下降,PaCO2、乳酸浓度、肺组织MPO活性、W/D比及TLR4表达和血清TNF-α浓度升高(P＜0.01);与ALI组比较,PHcD组pH值和PaO2升高,PaCO2、乳酸浓度、肺组织MPO活性、W/D比及TLR4表达和血清TNF-α浓度降低(P＜0.05).PHcD组肺组织病理性损伤较ALI组减轻.结论 盐酸戊乙奎醚可减轻大鼠胸部撞击诱发的急性肺损伤,其机制与下调肺组织TLR4表达,降低炎性反应有关.

Abstract：

Objective To investigate the effects of penehyclidine hydrochloride (PHCD) on acute lung injury (ALI) induced by blunt chest trauma and Toll-like receptor 4 (TLR4) expression in the lung tissues in rats.Methods Ninety-six male SD rats weighing 250-300 g were randomly divided into 3 groups ( n = 32 each):control group (group C), ALI group and PHCD group. ALI was induced by dropping a 300 g weight onto a precordial protective shield to direct the impact force away from the heart and toward the lungs in anesthetized rats according to the method described by Raghavendran et al. PHCD 2 mg/kg was injected intraperitoneally immediately after ALI was induced in group PHCD. Eight rats were selected at 2, 8, 12 and 24 h after ALI was induced, and arterial blood samples were collected for determination of the serum TNF-α concentration. Eight rats were selected at 8 h after ALI was induced, arterial blood samples collected for blood gas analysis and then the rats sacrificed. The lungs were immediately removed for determination of W/D lung weight ratio, myeloperoxidase (MPO) activity and TLR4 expression, and microscopic examination. Results The pH value and PaO2 were significantly lower, and the PaCO2, lactic acid level, MPO activity, W/D ratio, TLR4 expression and serum TNF-α concentration higher in groups ALI and PHCD than in group C (P ＜ 0.01 ). The pH value and PaO2 were significantly higher, and the PaCO2, lactic acid level, MPO activity, W/D ratio, TLR4 expression and serum TNF-α concentration lower in group PHCD than in group ALI ( P ＜ 0.05). The lung histopathologic damage was significantly ameliorated in PHCD group as compared with ALI group. Conclusion PHCD can protect the lungs against blunt chest trauma-induced ALI, and the down-regulation of TLR4 expression in lung tissues and reduction of inflammatory response are involved in the mechanism.     

创伤性急性肺损伤时兔肺微血管内皮细胞Cx40调控内皮细胞通透性的研究

目的 观察创伤性急性肺损伤(ALI)兔血清对肺微血管内皮细胞连接蛋白40(Cx40)表达和间隙连接通道(GJC)功能的影响,及后者与内皮细胞单层通透性的关系.方法 组织块贴壁法分离、培养兔肺微血管内皮细胞.细胞分为3组,对照组、创伤血清处理组和阻滞剂组.在阻滞剂组,细胞GJC阻滞剂1-庚醇(0.5 mmol/L)加入到培养液中.1 h后,阻滞剂组和创伤血清组的20%胎牛血清的培养液均被更换为含有20%创伤血清的培养液.对照组则只更换培养液.3组细胞继续孵育6 h后,进入实验观察.分别行划痕实验、Cx40蛋白免疫印迹、单层细胞通透性和细胞内Ca2+钙浓度检测.结果 Cx40蛋白印迹见创伤血清组与阻滞剂组蛋白表达水平较对照组有明显降低,且2组之间亦不同,阻滞剂组蛋白水平较创伤血清组低,吸光度和值减少(对照组726293.2±83 684.1,创伤血清组397 798.1±87 145.7,阻滞剂组316977.6±76 325.9,n=4,P＜0.05).3组细胞划痕边缘的细胞均被染料荧光黄(LY)染色,LY在对照组细胞扩散得最远,显色的周边细胞的数量最多,创伤血清组次之,阻滞剂组显色的细胞数量最少,相邻细胞与边缘细胞比值的差异有统计学意义(23±5、11±3比7±2,n=4,P＜0.05).3组细胞伊文思蓝清除率均随着时间的延长而增加,且3组细胞间亦有不同,阻滞剂组清除率高于创伤血清组和对照组.组间差异和时间点差异均有统计学意义(P＜0.05).通过共聚焦显微镜的观察,3组细胞荧光强度不相同,阻滞剂组荧光最强,对照组最弱.创伤血清组和阻滞剂组细胞内Ca2+浓度均显著高于对照组[(494.80±41.94)、(569.80±6.70)nmol/L比(158.80±13.09)nmol/L,P＜0.05].结论 创伤性ALI动物血清抑制肺微血管内皮细胞Cx40表达,进而引起GJC功能下降,导致肺血管内皮细胞单层通透性增加,该机制可能参与了胸部创伤后肺血管通透性增加的形成,促进了伤后急性肺损伤的发病.

Abstract：

Objective To investigate the influence of traumatic acute lung injury (ALI) rabbit serum on gap junction channel ( GJC ) function and connexin40 ( Cx40) expression, and the correlation of GJC and Cx40 with rabbit pulmonary microvascular endothelial cells permeability. Methods Cultured pulmonary microvessel endothelial cells were divided into three groups, control ( Gcontrol) , injured serum (Gserum) , and blocker agent ( Gblocker). GJC function was assessed by scrape-loading and dye transfer techniques. Pulmonary microvessel endothelial cells permeability was measured by Evans blue-labeled albumin transfer, and the expression of Cx40 was measured by Western blotting. Intracellular free calcium concentration was measured by fluo-3am. Results Injured serum decreased GJC function and the level of Cx40,which was aggravated by the GJC blocker (control 726 293. 2 ± 83 684. 1, serum 397 798. 1 ± 87 145. 7, blocker 316 977. 6 ±76 325. 9,n=4,P ＜0. 05). Similarly, pulmonary microvascular endothelial cells permeability was increased significantly in Gserum and Gblocker as compared with Gcontrol (P ＜0. 05). After treatment with injured serum in combination with gap junction blocker in vitro, intracellular free calcium concentration was increased [control (494. 80 ±41.94) nmol/L, serum (569. 80 ±6. 70) nmol/L, blocker (158. 80 ±13. 09) nmol/L, P＜ 0.05]. Conclusion Traumatic ALI rabbit serum inhibits the Cx40 expression and down-regulates GJC function, which contributes to the increase of pulmonary microvascular endothelial cells permeability after ALI This mechanism may participate in the increase of pulmonary vascular permeability and promote the morbidity of ALI after trauma.     

Proteinase 3 enhances endothelial monocyte chemoattractant protein-1 production and induces increased adhesion of neutrophils to endothelial cells by upregulating intercellular cell adhesion molecule-1

Wegener's granulomatosis is an autoimmune disease that is characterized by systemic vasculitis and granuloma formation. Early influx of polymorphonuclear neutrophils (PMN), followed at a later stage by mononuclear cells, contributes to the granulomatous inflammation. Previous studies have shown that proteinase 3 (PR3), the major autoantigen in Wegener's granulomatosis, specifically binds to endothelial cells and plays a possible role in activation of these cells by enhancing interleukin-8 production, thus providing a chemotactic and activating stimulus for PMN. The present study demonstrated that PR3 enhances the production of monocyte chemoattractant protein-1 (MCP-1) by human umbilical vein endothelial cells (HUVEC) in a dose- and time-dependent manner. The PR3-induced increase in MCP-1 production was demonstrated at both the protein and the mRNA levels and was chemotactic for monocytes. In addition, it was demonstrated that PR3 induces a dose- and time-dependent increase in the expression of intercellular adhesion molecule-1 (ICAM-1) as determined by fluorescence-activated cell sorter analysis. The PR3-induced increase in expression of ICAM-1 was also demonstrated at the mRNA level. PR3 induced a slight increase in vascular cell adhesion molecule-1 expression and had no effect on the expression of both P- and E-selectin. Incubation of HUVEC for 24 h in the presence of PR3 resulted in a significant increase in adhesion of PMN, which was reduced to baseline levels in the presence of blocking monoclonal antibody anti-ICAM-1 or anti-CD18 or a combination of both. Monocytes showed a slight but statistically not significant increase in adhesion. Incubation of HUVEC with PR3 for 4 h did not result in enhanced adhesion of either PMN or monocytes. It was hypothesized that PR3, which may be released locally at inflammatory sites after activation of cytokine primed PMN, plays a role in endothelial cell activation by enhancing both interleukin-8 and MCP-1 production, thus providing a chemotactic and activating stimulus for both PMN and monocytes. In addition, PR3 may contribute to the ongoing inflammation by enhancing the adhesion of PMN to endothelial cells by upregulating ICAM-1 expression.     

核因子κB在烧伤血清诱导内皮细胞细胞间黏附分子1表达中的作用

目的探讨核因子κB(NF-κB)在烧伤血清诱导内皮细胞(EC)分泌细胞间黏附分子(ICAM)1中的作用。方法人脐静脉内皮细胞(HUVEC)培养后,分别用正常人血清(对照组)、烧伤患者血清(烧伤血清组)、吡咯烷二硫代氨基甲酸盐(PDTC)+烧伤患者血清(PDTC组)刺激。于刺激0.5、1.0、2.0、4．0 b时采用电泳迁移率分析法测定HUVEC NF-κB的活性;流式细胞术检测刺激3．0、6.0、12．0、24．0 h时HUVEC膜表面ICAM-1的表达。结果刺激后烧伤血清组、PDTC组细胞NF-κB活性均明显高于对照组(P<0．01),1．0 h时达峰值[(21.03±4．87)、(7.44±0．60)×104积分灰度值],以后逐渐降低;PDTC组明显低于烧伤血清组(P<0.01)。刺激后烧伤血清组、PDTC组ICAM-1的表达均增加,刺激12．0 h吋达峰值(平均荧光密度各为327±37、142±31),与对照组比较,差异有统计学意义(P<0．01);PDTC组刺激12．0、24．0 h时低于烧伤血清组(P<0．01)。结论烧伤血清通过活化NF-κB,从而启动EC对黏附分子的合成和释放,提示NF-κB在烧伤血清诱导EC分泌黏附分子过程中起重要作用。