双重抗血小板对脓毒症大鼠急性肾损伤保护作用及机制的研究

Protective effects and mechanism of aspirin combined with clopidogrel on acute kidney injury induced by sepsis

目的探讨脓毒症大鼠急性肾损伤（AKI）与血小板活化的关系及阿司匹林联合氯吡格雷双重抗血小板的效应。方法40只雄性Wistar大鼠随机分为对照组（N组）、内毒素组（LPS组）、阿司匹林组（A组）、氯吡格雷组（C组）、阿司匹林＋氯吡格雷组（A＋C组），每组8只。N组、LPS组均给予生理盐水1mL，A组、C组、A＋C组则分别给予阿司匹林（100mg／kg）、氯吡格雷（200mg／kg）、阿司匹林（100mg／ks）＋氯吡格雷（200mg／kg）灌胃3d预处理后，N组腹腔注射生理盐水（10mL／kg），余组腹腔注射LPS10mg／kg造模，造模6h后采集标本进行分析。检测血清胱抑素c（Cysc）、血肌酐（SCr）及尿素氮（BUN）；流式细胞仪测全血中血小板CD62p、TLR4表达水平、全血电阻法测血小板聚集率；ELISA法测血清TNF-α及IL-6水平；肾脏湿／干质量比（W／D）进行比较，取肾组织石蜡包埋切片，HE染色，光镜下观察大鼠肾脏组织结构改变。结果与LPS组比较，药物干预各组肾脏W／D、肾脏病理改变减轻；SCr、BUN、CysC、血小板聚集率、CD62p、TLR4表达水平及血清TNF-α、IL-6均有所降低（P〈0．05），而A＋C组下降最明显（P〈0．01），差异均有统计学意义。结论阿司匹林联合氯吡格雷对脓毒症大鼠AKI有协同保护作用，其机制可能为阻断血小板活化和聚集信号通路，降低血清TNF-α、IL-6水平，进而调节炎症反应。

Objective To investigate the possible protective effects of aspirin combined clopidogrel on acute kidney injury in rats with sepsis, and its possible mechanism. Methods Forty male Wistar rats were randomly divided into five groups： control group （group N）, endotoxin group （ group LPS）, aspirin group （ group A）, clopidogrel group （ group C ）, aspirin plus clopidogrel group （ group A ＋ C）, with 8 rats in each group. The rats in group N and group LPS were respectively treated with 1 mL normal saline, while the rats in groups A, C and A ＋ Cwere respectively given aspirin （ 100 mg/kg）, clopiogrel （200 mg/kg）, aspirin （100 mg/kg） ＋ clopidogrel （200 mg/kg） pretreatment by gavage for three days. Then normal saline was injected in control group （10 mL/kg） by intrapertoneal, rats in all the tested groups were administered with LPS（ 10 mg/kg） by the same way. The blood Cr, BUN, Cys C, TNF - α, IL - 6 and the expression level of platelet CD62p, TLR4, rate of platelet aggregation 6 h after LPS or saline injection were determined. At last, kidney wet/dry weight ratio （W/ D）, pathological changes in the kidney were observed by hematoxylin eosin （HE） staining. Results Compared with LPS model group, renal function, the level of Cys C, TNF - α and IL - 6 in serum, the expression of platelet aggregation, platelet CD62p, TLR4 and kidney W/D ratio, significantly decreasedin drug intervention groups. The histological changes also lessened, especially in drug combination group. Conclusion Aspirin and ctopidogrel can block the platelet activation pathway and inhibit the levels of TNF - α and IL - 6 in serum, thus modify the inflammatory response and provide synergetic effects against acute kidney injury induced by sepsis.