NOD2受体在抗β_(2)GPⅠ抗体诱导血小板炎性反应促血栓形成中的作用研究  

Role of NOD2 receptor for promoting thrombosis in platelet inflammatory response induced by anti-β_(2)GPⅠantibody

作　　者：骆杰 查才军 张平[3] 王兆鑫 吴言[4] 张文静[2] LUO Jie;ZHA Caijun;ZHANG Ping;WANG Zhaoxin;WU Yan;ZHANG Wenjing(Department of Laboratory Medicine,Affiliated Tenth People′s Hospital,Tongji University,Shanghai 200072,China;Department of Laboratory Medicine,Second Affiliated Hospital of Harbin Medical University,Harbin,Heilongjiang 150086,China;Department of Neurosurgery,Second Affiliated Hospital of Harbin Medical University,Harbin,Heilongjiang 150086,China;Department of Ultrasound Medicine,Second Affiliated Hospital of Harbin Medical University,Harbin,Heilongjiang 150086,China)

机构地区：[1]同济大学附属第十人民医院检验医学科,上海200072 [2]哈尔滨医科大学附属第二医院:检验医学科,黑龙江哈尔滨150086 [3]哈尔滨医科大学附属第二医院:神经外科,黑龙江哈尔滨150086 [4]哈尔滨医科大学附属第二医院:超声医学科,黑龙江哈尔滨150086

出　　处：《国际检验医学杂志》2022年第4期398-403,共6页International Journal of Laboratory Medicine

基　　金：国家自然科学基金青年基金项目(82001715);黑龙江省卫生和计划生育委员会科研课题(2018286);黑龙江省省属高校基本科研业务费科研项目(2017LCZX67)。

摘　　要：目的明确核苷酸结合寡聚域2(NOD2)受体在抗β_(2)糖蛋白Ⅰ抗体(anti-β_(2)GPⅠ)诱导血小板炎性反应,进而促血栓形成中的作用机制,探讨以NOD2受体为干预靶点对anti-β_(2)GPⅠ诱导的相关血栓形成机制的影响。方法选取2018年1月至2021年3月哈尔滨医科大学附属第二医院经CT、MRI或动静脉造影诊断为血栓且anti-β_(2)GPⅠ阳性的患者69例为血栓组;选取同期体检健康者78例作为健康对照组。分别提取血栓患者、健康对照者的血小板和血浆,其中健康对照者血小板需与anti-β_(2)GPⅠ/β_(2)GPⅠ共孵育。利用实时定量PCR(RT-PCR)检测NOD2受体mRNA表达;ELISA检测白细胞介素(IL)-1β水平;免疫印迹法(Western blot)检测Toll样受体4(TLR4)、NOD2受体、p38丝裂原活化蛋白激酶(MAPK)活化情况;FeCl3诱导小鼠颈总动脉血栓形成,利用激光散斑超声血流仪监测血管闭塞时间。结果与健康对照组相比,血栓组患者血小板NOD2受体mRNA表达和血浆IL-1β水平显著增加(P<0.05);anti-β_(2)GPⅠ/β_(2)GPⅠ刺激后血小板NOD2受体mRNA表达增加,刺激后上清液IL-1β水平显著升高,差异均有统计学意义(P<0.05)。与anti-β_(2)GPⅠ/β_(2)GPⅠ刺激后相比,加入p38 MAPK抑制剂SB203580后,IL-1β水平显著降低;TLR4抑制剂TAK242可部分抑制NOD2受体蛋白表达,显著抑制p38 MAPK磷酸化和IL-1β水平;姜黄素可使p38 MAPK磷酸化及IL-1β水平下降;向小鼠体内注射姜黄素可使anti-β_(2)GPⅠ/β_(2)GPⅠ诱导的血栓形成时间显著延长。结论anti-β_(2)GPⅠ可通过TLR4调控NOD2受体/p38 MAPK信号轴影响IL-1β释放,姜黄素可通过抑制NOD2受体表达,降低p38 MAPK磷酸化及IL-1β水平,减缓anti-β_(2)GPⅠ诱导的炎性反应和血栓形成,有望为anti-β_(2)GPⅠ阳性血栓患者治疗提供新思路。Objective To clarify the mechanism of nucleotide-binding oligomerization-2(NOD2)receptor in anti-β_(2) glycoproteinⅠantibody(anti-β_(2)GPⅠ)-induced platelet inflammatory and thus promoting thrombosis,and to explore the effect of NOD2 receptor as an intervention target on anti-β_(2)GPⅠinduced related thrombosis.Methods Sixty-nine patients with thrombus and anti-β_(2)GPⅠpositive diagnosed by CT,MRI or arteriovenous angiography in the Second Affiliated Hospital of Harbin Medical University from January 2018 to March 2021 were selected as the thrombus group.Contemporaneous healthy subjects undergoing physical examination were selected as the healthy control group.Platelet and plasma were extracted from the patients with thrombus and healthy controls,and platelets from healthy controls were co-incubated with anti-β_(2)GPⅠ/β_(2)GPⅠ.The real-time PCR was used to detect the expression of NOD2 receptor mRNA.The IL-1βlevel was detected by ELISA.Toll-like receptor 4(TLR4),NOD2 receptor and p38 mitogen-activated protein kinase activation(MAPK)were detected by Western blot.FeCl_(3) induced the formation of common carotid artery thrombosis in mice,and the time of vascular occlusion was monitored by laser speckled ultrasonic flowmeter.Results Compared with the healthy control group,the platelet NOD2 receptor mRNA expression and plasma IL-1βlevel in the thrombus group were significantly increased(P<0.05).After anti-β_(2)GPⅠ/β_(2)GPⅠstimulation,the expression of platelet NOD2 receptor mRNA was increased,the level of IL-1βin stimulated supernatant was significantly increased,and the differences were statistically significant(P<0.05).Compared with the anti-β_(2)GPⅠ/β_(2)GPⅠstimulation,the level of IL-1βwas significantly decreased after the adding the p38 MAPK inhibitor SB203580.TLR4 inhibitor TAK242 could partially inhibit NOD2 receptor protein expression,significantly inhibited the p38 MAPK phosphorylation and IL-1βlevel.Curcumin could decrease p38 MAPK phosphorylation and IL-1βlevel.Injecting c

关 键 词：核苷酸结合寡聚域2受体 抗β_(2)糖蛋白Ⅰ抗体 炎性反应 血栓 姜黄素 

分 类 号：R446.1[医药卫生—诊断学]