Changes of L-selectin expression on peripheral blood polymorphonuclear leukocytes and their significance in rats with acute lung injury

AIM:To explore the changes of L-selectin expression on peripheral blood polymorphonuclear leukocytes (PMNs) and their significance in rats with acute lung injury (ALI). METHODS:ALI model in rat was established by intravonous injection of E. coli endotoxin (ET). The expression of L-selectin on peripheral blood PMNs was measured by immunofluorescence and flow cytometry.RESULTS:The contribution of L-selectin fluorescence signal was on the surface of PMNs membrane. The expression of L-selectin on poeripheral blood PMN was significantly lower at 5 min after injection of ET and the lowest during 15 min to 30 min, then gradually increased, but the expression of L-selectin on PMN was lower at 60 min after injection of ET than that of control animal.CONCLUSION:In physiological state, L-selectin were expressed on the surface of PMN membrane. The protein expression of L-selectin on PMNs reduced rapidly after injection of ET and the lowest at 15 min, then gradually increased. L-selectin may play a role in the development of ALI.     

L-选择素分子结构及功能的研究进展

近 1 0年来 ,由于分子生物学及相关学科的理论和技术的飞速发展 ,使得对L 选择素的分子结构、分子功能的研究更加深入。就其分子结构而言目前已经完全清楚 ,而功能与其相应的问题仍在研究和探索之中。作为受体的选择素 ,其功能的实现是在其与相应的配体结合之后。表达与调节是在细胞因子、炎症介质以及其他因素的作用下 ,细胞表面黏附分子表达的水平和构型可以发生改变 ,以调节免疫平衡。L 选择素通过白细胞表面的Ca2 依赖的凝集素结构域和胞内小片段激活酪氨酸酶p56lck和ras的信号途径进行跨膜信号转导。活化依赖和活化独立的脱落、生理和病理意义及基因改造对功能的影响等方面目前仍在进一步的研究之中     

乳牛L-选择素单克隆抗体的制备与亚类鉴定

以乳牛L-选择素为免疫抗原免疫BALB／c小鼠1只，相隔14d进行3次免疫，抗原用量每次30big，首次免疫后第116d进行尾静脉加强免疫，抗原用量10μg。免疫脾细胞与骨髓瘤细胞融合后接种12块96孔板。以L-选择素包被酶标板，间接ELISA检测阳性孔，有限稀释法克隆阳性孔，筛选出6株杂交瘤细胞。小鼠腹腔注射杂交瘤细胞株制备McAb，并对其分别进行Ig亚类测定。结果，6株杂交瘤细胞7810、10F6、10E2、5C6、12G7和llA3的Ig亚类分别是IgG2a、IgG2a、IgG2b、IgG1、IgG1和IgG1。     

乳牛L-选择素在中性粒细胞中表达与分布的定位研究

采用间接荧光标记及胶体金标记方法通过流式细胞仪和低压扫描电子显微镜，检测了L-选择素在健康泌乳中期乳牛（A组）及患金黄色葡萄球菌型临床型（B组）和亚临床型乳腺炎乳牛（C组）中性粒细胞上表达的基本规律。结果表明：三者表达L-选择素的中性粒细胞阳性率分别为97．90％、97．69％和96．67％（收集5000个细胞），组间差异不显著（P〉0．05），但患亚临床型乳腺炎乳牛的阳性率从3次测定的结果看是逐渐增加的；L-选择素表达的间接荧光强度组间差异极显著（P〈0．01）；而患亚临床型乳腺炎的乳牛组内差异也极显著（P〈0．01）；扫描电镜观察到的中性粒细胞表面脊样结构的分布及形态与流式细胞仪检测的结果具有相似性。     

Oxidative response of neutrophils to platelet-activating factor is altered during acute ruminal acidosis induced by oligofructose in heifers

Reactive oxygen species (ROS) production is one of the main mechanisms used to kill microbes during innate immune response. D-lactic acid, which is augmented during acute ruminal acidosis, reduces platelet activating factor (PAF)-induced ROS production and L-selectin shedding in bovine neutrophils in vitro. This study was conducted to investigate whether acute ruminal acidosis induced by acute oligofructose overload in heifers interferes with ROS production and L-selectin shedding in blood neutrophils. Blood neutrophils and plasma were obtained by jugular venipuncture, while ruminal samples were collected using rumenocentesis. Lactic acid from plasma and ruminal samples was measured by HPLC. PAF-induced ROS production and L-selectin shedding were measured in vitro in bovine neutrophils by a luminol chemiluminescence assay and flow cytometry, respectively. A significant increase in ruminal and plasma lactic acid was recorded in these animals. Specifically, a decrease in PAF-induced ROS production was observed 8 h after oligofructose overload, and this was sustained until 48 h post oligofructose overload. A reduction in PAF-induced L-selectin shedding was observed at 16 h and 32 h post oligofructose overload. Overall, the results indicated that neutrophil PAF responses were altered in heifers with ruminal acidosis, suggesting a potential dysfunction of the innate immune response.