摘要
目的:研究阿米福汀对苯并[a]芘(Ba P)诱导C57BL/6J小鼠腹主动脉瘤(AAA)形成的作用,并探讨相关机制。方法:体外培养RAW264.7单核巨噬细胞,分为正常对照组,溶剂对照组(即DMSO组),Ba P组,低剂量(1μmol/L)阿米福汀组,中剂量(5μmol/L)阿米福汀组及高剂量(25μmol/L)阿米福汀组,应用Western blot法检测体外培养的RAW264.7单核巨噬细胞基质金属蛋白酶(MMP)-9、MMP-12、TNF-α、NF-κB表达。48只C57BL/6J小鼠(8月龄,雄性)随机分为对照组、模型组(AngⅡ+Ba P组)、低剂量(50 mg/kg)阿米福汀组、高剂量(100 mg/kg)阿米福汀组。6周后取腹主动脉察看标本形态;行HE、Masson染色观察血管形态结构;测定腹主动脉的血管周长。Western blot、免疫组化法评价腹主动脉组织中的巨噬细胞浸润情况及MMP-9、MMP-12、TNF-α、NF-κB的表达。结果:Western blot发现Ba P刺激使巨噬细胞MMP-9、MMP-12、TNF-α、NF-κB的表达升高,而阿米福汀对此有显著抑制作用,且呈剂量依赖性(P<0.05)。动物实验可见对照组成瘤率为0,高剂量阿米福汀组成瘤率为16.67%,与模型组(58.33%)和低剂量阿米福汀组(33.33%)相比显著降低(P<0.05)。免疫组化结果显示高剂量阿米福汀组腹主动脉壁的巨噬细胞浸润程度及TNF-α、MMP-9、MMP-12、NF-κB的表达较模型组和低剂量阿米福汀组相比显著下降(P<0.05)。Western blot实验结果证实高剂量阿米福汀组腹主动脉壁TNF-α、MMP-9、MMP-12、NF-κB的表达较模型组和低剂量阿米福汀组相比显著下降(P<0.05)。结论:阿米福汀可以抑制Ba P诱导的巨噬细胞的活化,同时可以抑制Ba P诱导的小鼠腹主动脉瘤发生、发展,其机制可能跟抑制NF-κB途径、巨噬细胞浸润及MMPs、TNF-α的表达有关。
AIM：To study the role of amifostine on the formation of benzo[a]pyrene（ Ba P）-induced abdominal aortic aneurysm（ AAA） in C57 BL/6J mice and the underlying mechanism.METHODS：RAW246.7 mononuclear macrophage in vitro were divided into control group,DMSO group,Ba P group,low dose（ 1 μmol/L） amfostine treated group,middle dose（ 5 μmol/L） amfostine treated group and high dose（ 25μmol/L） amfostine treated group.The influence of Ba P on the expression of matrix metalloproteinase（ MMP）-9,MMP-12,TNF-α,NF-κB in the RAW246.7 mononuclear macrophages in vitro was determined by Western blot.Male C57 BL/6J mice（ 8 months old） were divided into control group,model group（ Ang II ＋ Ba P group）,low dose（ 50 mg/kg） amfostine treated group and high dose（ 100 mg/kg）amfostine treated group.After 6 weeks,the abdominal aorta were isolated.The aortic tissues were subjected to HE and Masson staining.The vascular wall structure,infiltration of macrophage,the expression of MMP-9,MMP-12,TNF-α,NF-κB were evaluated by Western blot and immunochemistry staining.RESULTS：Amifostine attenuated Ba P-induced expression of TNF-α,MMP-9,MMP-12,NF-κB in the RAW246.7 mononuclear macrophages（ P〈0.05）.The results of animal experiments showed that the incidence of AAA in high dose amifostine treated group were significantly lower than that in low dose amifostine treated group and model group（ P〈0.05）.Immunohistochemistry staining observation showed that amifostine inhibited the aortic macrophage infiltration more obviously in high amifostine treated group compared with model group and low dose amifostine treated group（ P〈0.05）.Compared with model group and low dose amifostine treated group,the MMP-9,MMP-12,TNF-α and NF-κB expression of abdominal aorta in high amifostine treated group was reduced significantly（ P〈0.05）.CONCLUSION：Amifostine inhibits Ba P-induced activation of macrophages,and also prevents the formation of abdominal aortic aneurysm in C57 BL/6J mice induced by Ba P by inhibition of the NF-κB pathway,macrophage infiltration and the expression of TNF-α and MMPs.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2016年第12期2168-2176,共9页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.81573185)
浙江省科技厅项目(No.2014C33163)
温州科技局项目(No.Y20140678)
作者简介
通讯作者Tel：0577-88002214;E-mail：jikt@wzmc.edu.cn
