白介素8对兔腹主动脉粥样硬化狭窄的影响

1、白介素8对兔腹主动脉粥样硬化狭窄的影响         10-09-10 10:04:00     编辑：studa20                         作者：齐晓勇 孟存良 李英肖 党懿 刘惠良   &

2、#160;                     王天红 袁华兵 倪艳辉 刘 光 武振朝 郜利会【摘要】  目的 观察白介素8（IL8）水平对兔腹主动脉粥样硬化狭窄的影响。方法 新西兰大白兔36只随机分为实验组、IL8单克隆抗体治疗组、对照组。实验组和治疗组动物通过穿刺股动脉送入球囊拉伤腹主动脉。治疗组动物通过耳缘静脉注射IL8单克隆抗体进行干预。对照组动物只进行股动脉穿刺和留置鞘管，

3、不进行球囊扩张拉伤。分别于实验前,实验后4 h、1 d、3 d，1 w、2 w、4 w，耳缘静脉采血4 ml，检测三组动物血清IL8水平，4 w后处死动物进行病理组织学检查，应用光学显微镜和计算机病理图像分析系统分析测定以下指标：内膜中膜厚度、面积，计算血管狭窄程度。结果 实验组动物球囊损伤4 h后IL8水平即开始升高，术后1 d达到峰值，持续增高4 w，且明显高于治疗组和对照组。治疗组和对照组动物IL8水平无上述变化规律。实验组动物腹主动脉明显狭窄，组织学检查内膜面积和中膜面积、血管狭窄程度等指标大于治疗组和对照组（P<0.01），治疗组管腔狭窄，对照组动物管腔无明显变化，治疗组与对照

4、组比较无显著性差异(P>0.05)。相关分析显示，IL8水平与内膜厚度、内膜面积、中膜厚度、中膜面积等呈正相关(相关系数r分别为0.894，0.783，0.801，0.912，均P<0.01)。结论 IL8水平高表达引起腹主动脉粥样硬化狭窄的产生，经IL8单克隆抗体治疗后动脉狭窄程度明显减轻。IL8水平与动脉狭窄程度呈正相关，是发生腹主动脉粥样硬化狭窄的独立危险因素。 【关键词】  腹主动脉；动脉粥样硬化；炎性反应；白介素8；单克隆抗体    【Abstract】  Objective  To observe the ef

5、fects of interleukin8 (IL8) on atherosclerosis(AS) of abdominal aorta in rabbits. Methods  Thirtysix New Zealand white rabbits were randomly divided into experiment, treatment and control groups, each group 12 cases. Rabbits in experiment and treatment groups were injuried in abdominal aorta by

6、 balloon inflation after punctured in femoral artery. Monoclonal antibody of IL8 was injected by venous in rabbits of treatment group. Rabbits in control group were punctured only in femoral artery. Peripheral blood was collected before experiment and 4 h, 1, 3 d, 1, 2, 4 w later. The level of IL8 w

7、as measured by enzyme linked immunosorbent assay (ELISA) respectively. The luminals of abdomial aorta in three groups were observed 4 w later. Histopathologic examination was performed. Such contents as follows were assayed by light microscope and computer image analysis system: luminal area, intima

8、 and tunica media area, angiostenosis of blood vessel. Results  The level of IL8 was raised in 4 h and achieved to peak in 1 d after balloon inflation in rabbits of experiment group. The higher level of this inflammatory factor lasted 4 w. There was no variation above in treatment and control g

9、roups. It was observed that abdominal aorta became stenosis obviously in experiment group. The area of intima and tunica media as well as the extent of stenosis in experiment group were bigger than those in treatment and control groups. There was no statistical difference between treatment and contr

10、ol groups. Correlation analysis indicated that there were positive relations between IL，0.783，0.801，0.912，P<0.01). Multiple regression analysis of Logistic indicated that IL8 was an independent risk for the AS（P<0.01）. The high levels of IL8 leaded to AS in abdominal aorta of rabbits. The seve

11、rity of stenosis became lessen after intervention by monoclonal antibody of IL8. Positive corelation was found between the level of IL8 and severity of arterial stenosis. IL8 was one of independent risks factor leading occurrence and development of AS. Conclusions  The high levels of IL8 leads

12、to AS in abdominal aorta of rabbits. The severity of stenosis becomes lessen after intervention by monoclonal antibody of IL8. Positive corelation is found between the level of IL8 and severity of arterial stenosis. IL8 is one of independent risks factor leading occurrence and development of AS.

13、0;   【Key words】  Abdominal aorta; Atherosclerosis; Inflammatory reaction; Interleukin8; Monoclonal antibody目前研究已经证实，炎性反应在动脉粥样硬化（AS）的发生与发展中起到重要的作用。参与AS形成的炎性因子众多，且互相影响，因此观察炎性因子的变化规律，了解炎性因子与AS病变的关系与作用机制，有助于寻找临床干预AS的新方法。白细胞介素8（Interleukin8，IL8）是体内重要的炎性因子，可促使血液中单核细胞聚集到内膜上及向内膜下转移。本研究拟采用IL8

14、单克隆抗体进行干预治疗，观察干预治疗后IL8水平变化以及对腹主动脉AS狭窄的影响，为临床治疗AS性疾病提供新思路。1  材料与方法1.1  动物与分组选取812月龄新西兰大白兔36只，雌雄不拘，体质量2.503.25 kg，由河北医科大学实验动物中心提供(许可证号：SCXK(冀)20031003)。以上实验动物随机分为球囊拉伤组（实验组,n=12）和IL8单克隆抗体干预组（治疗组,n=12）以及对照组（n=12）。1.2  方法1.2.1  兔腹主动脉AS模型的建立 实验动物术前12 h禁食，不禁水，称体重。常规建立耳缘静脉输液通路，以10 g

15、/L戊巴比妥钠3.03.5 ml/kg耳缘静脉注射麻醉实验动物，麻醉后动物仰卧固定于手术台，以20%硫化钠双侧腹股沟脱毛。常规消毒铺巾后，seldinger技术穿刺股动脉并留置鞘管，经鞘管注入普通肝素200单位抗凝，沿导引钢丝逆行插入球囊，连接手推式压力表，造影后将球囊定位，置于髂动脉以上2个椎体处，以405.30607.95 kPa压力扩张球囊，同时注射造影剂证实球囊完全扩张堵闭腹主动脉，自近心端缓慢回拉(拉伤长度4 cm)，反复3次，每次球囊扩张持续时间20 s，每次间隔15 s，撤出球囊，拔除动脉鞘管，压迫止血15 min后，绷带包扎固定，4 h后解除绷带包扎。1.2.2  I

16、L8单克隆抗体的干预治疗 干预组动物实验前开始通过耳缘静脉注射IL8单克隆抗体2 mg/kg，连续3 d。1.2.3  病理学检查 剥离腹主动脉大体标本（胸主动脉至髂动脉分叉处）。剪除血管外结缔组织及脂肪，生理盐水冲洗残血。截取成形部位及对照组血管标本，立即放入10%福尔马林溶液内浸泡固定。常规酒精脱水，石蜡包埋，制成3 m厚连续切片24 h，给予苏木素伊红（HE）染色。利用HE染色的切片，随机选取5个视野，在40倍光镜下，利用微机图像处理测量系统对切片进行图像摄取和转换，测量斑块纤维帽及新生内膜厚度（IT）、中膜厚度(MT)、内膜面积(IA)、中膜面积(MA)，

17、计算血管狭窄程度、IT/MT比值，以及IA/MA比值。1.3  统计学方法 采用SPSS13.0软件包完成统计处理，先进行组间均衡性检验，计量资料以x±s表示，采用2检验。先行方差分析，然后再作多组均数的q检验，相关资料采用相关分析法计算相关系数，并进行多元Logistic回归分析。2  结  果2.1  三组实验动物之间体重、月龄、性别、血脂等一般情况比较，差异无统计学意义（P>0.05），见表1。表1  三组动物一般情况比较(略)2.2  三组动物术前、术后IL8水平比较 术前三组IL8水平无明显差异；术后4 h实验组IL8水平较术前以及干预组和对照组明显升高，术后1 d达到峰值，持续增高4 w。干预组与治疗组无上述变化趋势，干预组显著低于实验组（P<0.01），与对照组比较，无显著性差异(P>0.05)，见表2。表2  三组之间不同时间点IL8水平(略)2.3  三组动物病理形态学改变  HE染色可见对照组动脉内膜光滑，内皮下有少量结缔组织，无平滑肌细胞存在，中膜平滑肌细胞成梭形，环绕管腔排列；实验组兔内膜、中膜明显增厚，管腔明显狭窄，内膜下平滑肌细胞明显增生，而且排列不规则，内皮及