CCK-8对内毒素血症小鼠腹腔巨噬细胞B7.1和B7.2表达及协同刺激功能的影响

目的： 探讨八肽胆囊收缩素（CCK-8）对内毒素血症小鼠腹腔巨噬细胞B7.1和B7.2表达及其协同刺激活性的影响。方法: 将BALB/c小鼠随机分组（n=4）,分别腹腔注射生理盐水（0.2-0.3 mL/mouse），LPS（100 μg/mouse）和/或CCK-8 (5 nmol/mouse)及CR1409（100 μg/mouse）、CR2945（100 μg/mouse）。12 h后收集并纯化腹腔巨噬细胞。采用流式细胞术分析细胞表面B7.1和B7.2含量的变化。用免疫磁珠从小鼠脾细胞分离CD4+T细胞，按4∶1数量比与上述处理的腹腔巨噬细胞共同体外培养，同时加入ConA 5 mg/L，采用［³H］掺入法测定CD4+T细胞增殖，反映巨噬细胞的协同刺激活性。结果: 整体应用CCK-8作用小鼠腹腔巨噬细胞，与对照组相比，CCK-8可上调静息小鼠腹腔巨噬细胞B7.2的表达，而对B7.1的表达则无影响；并且CCK-8使CD4+T细胞［³H］-TdR的掺入率升高，即促进其增殖，增强巨噬细胞的协同刺激活性。CCK-8降低LPS活化的内毒素血症小鼠腹腔巨噬细胞B7.1和B7.2的表达并且降低CD4+T细胞的［³H］-TdR掺入率，即抑制其增殖，抑制其协同刺激活性。CR1409及CR2945均能逆转CCK-8的上述作用，且CR1409的作用较CR2945更明显。结论: CCK-8通过上调巨噬细胞B7.2表达而增强其协同刺激活性；并且降低LPS活化的内毒素血症小鼠腹腔巨噬细胞B7.1和B7.2的表达，抑制其协同刺激活性。该作用由CCK1R及CCK2R共同介导，其中CCK1R起主要介导作用。     

大承气汤对内毒素血症小鼠血浆内皮素-1及肾功能的影响

目的观察中药大承气汤对内毒素血症血浆内皮素-1及肾功能的影响。方法66只小鼠均分为三组：对照组,内毒素血症组（腹腔注射内毒素,立即、4h用生理盐水灌胃）,和治疗组（腹腔注射内毒素,立即、4h用大承气汤灌胃）。注射4h、8h后眼眶取血测定血清血浆内皮素-1（ET-1）和血清肌酐。结果注射内毒素后4和6h,治疗组血浆内皮素一1水平分别为（363．669±31．747）pg／ml和（323．320±43．327）pg／ml,均显著低于内毒素血症组[（470．028±49．430）和（462．917±20．13）pg／ml,均P〈0．01]。注射内毒素后4和8h,治疗组血清肌酐水平分别为（40．1±3．71）μmol／L和（39．29±3．71）μmol／L,均显著低于内毒素血症组[（48．18±5．15）和（47．38±4．14）μmol／L,均P〈0．01]。结论大承气汤能降低内毒素血症血浆内皮素水平和血清肌酐水平,减轻肾脏损伤,其机制可能与抑制内皮素有关。     

Acute phase protein, α – 1- acid glycoprotein (AGP-1), has differential effects on TLR-2 and TLR-4 mediated responses

Alpha-1-acid glycoprotein (AGP-1) is a major positive acute phase glycoprotein with unknown functions that likely play a role in inflammation. We tested its involvement in a variety of inflammatory responses using human AGP-1 purified to apparent homogeneity and confirmed its identity by immunoblotting and mass spectrometry. AGP-1 alone upregulated MAPK signaling in murine peritoneal macrophages. However, when given in combination with TLR ligands, AGP-1 selectively augmented MAPK activation induced by ligands of TLR-2 (Braun lipoprotein) but not TLR-4 (lipopolysaccharide). In vivo treatment of AGP-1 in a murine model of sepsis with or without TLR-2 or TLR-4 ligands, selectively potentiated TLR-2-mediated mortality, but was without significant effect on TLR-4-mediated mortality. Furthermore, in vitro, AGP-1 selectively potentiated TLR-2 mediated adhesion of human primary immune cell, neutrophils. Hence, our studies highlight a new role for the acute phase protein AGP-1 in sepsis via its interaction with TLR-2 signaling mechanisms to selectively promote responsiveness to one of the two major gram-negative endotoxins, contributing to the complicated pathobiology of sepsis.     

Evaluation of the effects of a pneumoperitoneum with carbon dioxide or helium in a porcine model of endotoxemia

Background: The expansion of the laparoscopic techniques to the critically ill patient is currently under debate. In order to evaluate the potential risks of performing laparoscopy in a body with signs of sepsis, the effects of the pneumoperitoneum were studied in a porcine model of mild endotoxemia. Methods: Twenty-eight pigs were separated into four groups of seven animals: untreated control (C), 2 μg/kg/h endotoxin (E), endotoxin and a pneumoperitoneum (3 h, 15 mmHg) with CO₂ (EC) or with helium (EH). Hemodynamic and homeostatic variables were studied for a period of 5.5 h. Primary endpoints were arterial and mucosal pH and the ATP content of the bowel wall. Statistical evaluation was performed using analysis of variance and the Bonferroni test. Results: Endotoxin infusion induced characteristic symptoms of early sepsis: increase of arterial CO₂, pulmonary arterial, portal venous, and pulmonary artery wedge pressure, and decrease of arterial pressure, cardiac output, arterial and mucosal pH. An additional pneumoperitoneum led to aggravation of all criteria with significant alterations in arterial and mucosal pH, arterial CO₂, wedge and portal venous pressure. The most striking derangement of mean values was observed for mucosal pH (EC: 7.40, EH: 7.54) and arterial pH (EC: 7.15, EH: 7.18). In group EC, two animals died in septic shock. Conclusion: Applying a pneumoperitoneum during an ongoing sepsis significantly deteriorates hemodynamic and homeostatic variables, thus enhancing the risk of severe complications. Received: 26 July 1999 In revised form: 21 January 2000 Accepted: 24 January 2000     

卡巴胆碱对内毒素血症小鼠肠道屏障功能的影响

目的观察卡巴胆碱对内毒素血症小鼠肠道屏障功能的保护作用及机制。 方法将C57BL/6小鼠随机分为对照组、内毒素血症组、卡巴胆碱组、α银环蛇毒素组,每组10只。腹腔注射10 mg/kg内毒素建立内毒素血症模型。模型制备后15 min腹腔注射卡巴胆碱0.1 mg/kg或0.9%氯化钠溶液,内毒素注射后3 h处死动物。取距回盲瓣10 cm处回肠组织,光镜下观察回肠组织病理学改变,测定肠道组织通透性,检测紧密连接蛋白（Claudin-2）、肌球蛋白轻链激酶（MLCK）定位及蛋白含量等。 结果FITC-葡聚糖水平：对照组、内毒素血症组、卡巴胆碱组、α银环蛇毒素组分别为（2.33±0.51）μg/ml、（55.25±5.41）μg/ml、（19.27±3.53）μg/ml、（48.45±9.50）μg/ml,4组总体差异有统计学意义（F=111.8,P<0.05）,其中内毒素血症组与对照组、内毒素血症组与卡巴胆碱组、α银环蛇毒素组与卡巴胆碱组比较,差异均有统计学意义（t分别为22.52、15.31、12.42,P均<0.05）。Claudin-2蛋白含量：对照组、内毒素血症组、卡巴胆碱组、α银环蛇毒素组分别为（0.82±0.08）μg/ml、（0.52±0.09）μg/ml、（0.77±0.05）μg/ml、（0.53±0.09）μg/ml,4组总体差异有统计学意义（F=11.61,P<0.05）,其中内毒素血症组与对照组、内毒素血症组与卡巴胆碱组、α银环蛇毒素组与卡巴胆碱组比较,差异均有统计学意义（t分别为6.518、5.366、5.167,P均<0.05）。MLCK蛋白含量：对照组、内毒素血症组、卡巴胆碱组、α银环蛇毒素组分别为（0.58±0.07）μg/ml、（1.07±0.17）μg/ml、（0.69±0.11）μg/ml、（0.94±0.05）μg/ml,4组总体差异有统计学意义（F=12.64,P<0.05）,其中内毒素血症组与对照组、内毒素血症组与卡巴胆碱组、α银环蛇毒素组与卡巴胆碱组比较,差异均有统计学意义（t分别为7.79、5.881、3.892,P均<0.05）。 结论卡巴胆碱对内毒素血症小鼠肠道屏障功能有保护作用,可降低肠道通透性,其机制可能与激活胆碱能抗炎通路有关。     

内毒素血症对糖尿病大鼠细胞因子、HPA轴功能的影响

目的观察内毒素血症对糖尿病大鼠细胞因子、HPA轴功能的影响。方法用链脲佐菌素(STZ,50mg/kg)予SD大鼠腹腔注射建立糖尿病动物模型,在此基础上再以脂多糖[LPS,0.5mg/(kg bw)]腹腔注射诱导内毒素血症模型,观察注射LPS后6、12、24及72h各时间点血糖、胰岛素、TNF-α、IL-6及CRH、ACTH、皮质酮的变化。结果内毒素血症糖尿病大鼠血清TNF-α、IL-6水平显著升高,于注射LPS后12h达峰值(P<0.05);而DM+LPS各亚组血浆CRH、ACTH、皮质酮水平略有上升,各组间比较均无显著差异(P>0.05);各时间点的血糖及胰岛素水平变化无统计学意义。结论内毒素血症可使糖尿病大鼠细胞因子分泌增加,激活HPA轴功能。     

己酮可可碱对内毒素血症大鼠胃黏膜损伤的保护作用

目的探讨己酮可可碱（PTX）对内毒素血症大鼠胃黏膜损伤的保护作用。方法Wistar大鼠30只随机分为三组：假手术对照组、内毒素组和PTX治疗组,每组10只。检测胃黏膜细胞间黏附分子-1（ICAM-1）表达,并测定各组动物血清TNF—α、IL-1β水平。结果内毒素组ICAM．1表达与假手术组比较明显增高（P〈0．01）,其血清TNF-α和IL-1β水平与假手术组比较也明显增高（P〈0．01）。PTX治疗后,该组血清TNF-α、IL-1β水平和胃黏膜组织ICAM-1蛋白表达与内毒素组比较明显减弱（P〈0．01）。结论ICAM-1和TNF-α、IL-1β在内毒素急性胃黏膜损伤发病机制中起重要作用,PTX可以降低血循环TNF-α、IL-1β水平和下调胃黏膜组织中ICAM-1表达,减轻内毒素对胃黏膜上皮组织的损伤,保护胃黏膜。     

烫伤大鼠细胞免疫抑制与肠源性内毒素血症的关系

采用大鼠40%TBSAⅢ度烫伤模型,将动物随机分成烫伤对照组和选择性消化道脱污染(SI)D)防治组,探讨烫伤大鼠全身性细胞免疫抑制与肠源性内毒素血症的关系。结果显示,大鼠40%Ⅲ度烫伤后睥细胞对促有丝分裂原增殖应答反应、诱生白介素-2(IL-2)活性及 T 细胞亚群(Th/Ts)比值明显下降。预防性进行 SDD 动物,内毒素血症发生率显著降低,脾细胞增殖应答反应和 IL-2活性的诱生能力均明显恢复(P0.05)。提示烫伤所致肠道细菌/内毒素移位可能与诱发机体细胞免疫功能异常密切相关。SDD 预处理可防止动物细菌/内毒素移位,从而减轻创伤后的免疫抑制。     

血清新喋呤与烧伤病人内毒素血症及脓毒症的关系

为探讨血清新喋呤与大面积烧伤后内毒素血症及脓毒症的关系，对３５例烧伤面积大于３０％（３０％～９８％）患者的血清新喋呤、血浆内毒素的变化进行了动态观察。结果表明，烧伤后第３天大多数患者新喋呤升高（Ｐ＜０．０５），但与烧伤面积无显著相关（Ｐ＞０．０５）。严重烧伤第２周以后内毒素血症患者新喋呤水平显著高于无内毒素血症患者（Ｐ＜０．０５～０．０１）。同时，伤后第１４，２１天内毒素血症患者的循环内毒素与新喋呤呈显著正相关（第１４天：ｒ＝０．３６８，Ｐ＜０．０５；第２１天：ｒ＝０．４３９，Ｐ＜０．０１）。在整个监测期间，脓毒症患者（１５例）血清新喋呤水平持续升高，伤后第１４，２８天与非脓毒症患者相比差异有显著或非常显著意义（Ｐ＜０．０５～０．０１）。提示烧伤可引起血清新喋呤升高，但与烧伤面积无关。烧伤后内毒素血症对新喋呤的不断产生可能有显著影响。血清新喋呤的持续升高与严重烧伤后脓毒症的发生与发展密切相关。     

Extracorporeal Endotoxin Removal by Immobilized Polyethylenimine

Abstract: The neutralization of bacterial endotoxins (ET) is still an unsolved problem in therapeutic medicine. The efficacy of antiendotoxin antibodies or receptor antagonists and other substances interferring with the endotoxininduced pathomechanisms is dependent on an intact cellular degradation system of the host. However, the phagocytosis function of that system seems to be impaired regularly in patients with intense or long-lasting endotoxemia or septic shock and in patients undergoing hemodialysis. Extracorporeal adsorption of ET might well be an effective support in the anti-ET therapy by lowering the amount of circulating ET and thus relieving the defense system of the body. In this work a new ET-adsorbent based on macroporous cellulosic beads with immobilized polyethylenimine (PEI) was tested for its ET-removal capacity in vitro. A test solution with 100 ng/ml ET from Escherichia coli 055:B5 was recirculated in a system containing the adsorbent beads. Polymyxin B immobilized to the same carrier was used for comparison. PEI as well as polymyxin B showed complete removal of ET from plasma and water as was measured by the Limulus Amebocyte Lysate (LAL) test (Chromogenix). The biocompatibility of the PEI adsorber was superior to that of polymyxin B. The results indicate that the PEI adsorber is of high efficacy and possibly of interest for the treatment of endotoxemia.