黄芪桂枝五物汤防治深静脉血栓形成模型大鼠的实验研究

Experimental study of Huangqi Guizhi Wuwu Decoctionin prevention and treatment of rats model with deep venous thrombosis

  • 摘要:
    目的 探讨黄芪桂枝五物汤防治深静脉血栓形成(DVT)模型大鼠的作用机制。
    方法 将40只无特定病原体(SPF)级SD雄性大鼠随机分为假手术组、模型组、中药组及肝素钙组,每组10只。除假手术组外,其他3组均构建不完全结扎的大鼠股静脉血栓模型。中药组采用黄芪桂枝五物汤冻干粉溶液2.0 g/(kg·d)灌胃; 肝素钙组采用低分子量肝素钙351.43 IU/(kg·d)皮下注射; 假手术组、模型组均以蒸馏水灌胃; 各组大鼠给药时间均为造模前7 d至造模后7 d,给药1次/d,持续给药14 d。治疗后取各组大鼠腹主动脉血,检测活化部分凝血活酶时间(APTT)、凝血酶原时间(PT)、凝血酶时间(TT)、纤维蛋白原(FIB)、D-二聚体(D-D)等凝血指标,并进行血浆代谢产物分析; 取各组大鼠股静脉段,观察静脉血栓形成段的组织形态学差异。
    结果 与假手术组比较,模型组大鼠FIB及D-D水平均升高,APTT、PT、TT均缩短,差异有统计学意义(P < 0.05); 与模型组比较,肝素钙组、中药组FIB及D-D水平均下降,APTT、PT、TT均延长,差异有统计学意义(P < 0.05); 与肝素钙组比较,中药组D-D水平更低,差异有统计学意义(P < 0.05)。血浆代谢产物分析显示,模型组与假手术组共筛选出27种差异代谢物; 黄芪桂枝五物汤可以使14种差异代谢物显著回调,主要涉及改善维生素K、三甲基胺氮氧化物(TMAO)、鹅肌肽、L-肌肽、肉碱代谢、胆汁酸代谢和脂代谢等; 低分子量肝素钙可以使8种差异代谢物显著回调,主要涉及TMAO、鹅肌肽、L-肌肽、肉碱代谢以及氨基酸代谢等。组织形态学结果显示,假手术组静脉管腔中无血栓形成; 模型组中静脉管腔被血栓填塞,炎性细胞浸润较为严重; 肝素钙组静脉管腔内部分血栓崩解,部分血栓收缩,炎性细胞浸润较模型组轻; 中药组静脉管腔内血栓大部分崩解消失,残留少量血栓收缩,管壁炎性细胞浸润较轻,提示病理损害程度较轻。
    结论 黄芪桂枝五物汤具有抗凝、改善代谢紊乱、抑制血栓形成、保护血管内皮细胞以及减轻炎性细胞浸润等作用,这可能是其治疗DVT的重要机制。

     

    Abstract:
    Objective To investigate the mechanism of Huangqi Guizhi Wuwu Decoction in prevention and treatment of rats model with deep venous thrombosis (DVT).
    Methods A total of 40 male SD rats with specific pathogen free (SPF) grade were randomly divided into sham operation group,model group,Chinese medicine group and heparin calcium group,with 10 rats in each group. Except for the sham operation group,the femoral vein thrombosis models of rats with incomplete ligation were established in the other three groups. In the Chinese medicine group,2.0 g/(kg·d) of freeze-dried powder solution of Huangqi Guizhi Wuwu Decoction was given by intragastric administration; in the heparin calcium group,351.43 IU/(kg·d) of low molecular weight heparin calcium was given by subcutaneous injection; in the sham operation group and the model group,distilled water was given by intragastric administration; the administration time for rats in each group was from 7 days before modeling to 7 days after modeling,once a day,lasting for 14 days. After treatment,the abdominal aorta blood of rats in each group was collected to detect the coagulation indicators such as activated partial thromboplastin time (APTT),prothrombin time (PT),thrombin time (TT),fibrinogen (FIB) and D-dimer (D-D),and the plasma metabolites were analyzed; the femoral vein segments of rats in each group were collected to observe the histomorphological differences of venous thrombosis segments.
    Results Compared with the sham operation group,the levels of FIB and D-D in the model group increased significantly,and APTT,PT and TT shortened significantly (P < 0.05); compared with the model group,the FIB and D-D levels decreased significantly in the heparin calcium group and the Chinese medicine group,while APTT,PT and TT prolonged significantly (P < 0.05); compared with the heparin calcium group,the level of D-D in the Chinese medicine group was significantly lower (P < 0.05). Plasma metabolic analysis showed that there were 27 differential metabolites between the model group and the sham operation group; Huangqi Guizhi Wuwu Decoction could significantly restore 14 differential metabolites,involving the improvement of vitamin K,trimethylamine N-oxide (TMAO),anserine,L-carnosine,carnitine metabolism,bile acid metabolism and lipid metabolism; low molecular weight heparin could significantly restore 8 differential metabolites,involving TMAO,anserine,L-carnosine,carnitine metabolism and amino acid metabolism. Histomorphological results showed that there was no thrombosis in the venous lumen in the sham operation group; in the model group,the venous lumen was filled with thrombus,and the inflammatory cell infiltration was more serious; in the heparin calcium group,part of the thrombus in the venous lumen disintegrated,part of the thrombus contracted,and the inflammatory cell infiltration was lighter than that in the model group; in the Chinese medicine group,most of the thrombus in the venous lumen disintegrated and disappeared,a small amount of thrombus remained and contracted,and the inflammatory cell infiltration in the vascular wall was light,which indicated the degree of pathological damage was light.
    Conclusion Huangqi Guizhi Wuwu Decoction has the effects of anticoagulation,improving metabolic disorder,inhibiting thrombosis,protecting vascular endothelial cells and reducing inflammatory cell infiltration,which may be the important mechanism for treatment of DVT.

     

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