化疗对大鼠肺癌模型凝血功能及血管内皮功能的影响及机制研究

Effects and mechanism of chemotherapy on coagulation function and vascular endothelial function in rat lung cancer model

目的:观察化疗对大鼠肺癌模型凝血功能和血管内皮功能的影响,并分析其作用机制。方法:选取30只SD大鼠,随机分为正常组、模型组和化疗组,每组10只。模型组和化疗组采用致癌碘油液气管滴注方法建立大鼠肺癌模型,正常组大鼠仅滴注碘油液。化疗组大鼠采用5 mg/kg环磷酰胺腹腔注射,每天1次,每周5天,连续8周,正常组和模型组大鼠注射等量生理盐水。最后一次给药2 h后检测大鼠血浆活化部分凝血活酶时间(APTT)、凝血酶原时间(PT)、凝血酶时间(TT)和纤维蛋白原(FIB)含量,ELISA法检测血浆内皮素(ET-1)、一氧化氮(NO)和血管内皮生长因子(VEGF)的含量,Western blot法检测主动脉内皮Toll样受体4(TLR4)和NF-κB p65的表达水平。结果:与正常组比较,模型组大鼠APTT、PT、TT缩短,FIB含量增加;与模型组比较,化疗组大鼠APTT、PT、TT进一步缩短,FIB含量亦增加。ELISA检测结果显示,正常组、模型组和化疗组大鼠血浆中ET-1的含量分别为(35.67±3.89) ng/L、(51.61±4.14) ng/L、(59.57±4.51) ng/L;NO的含量分别为(90.31±10.61)μmol/L、(124.92±13.26)μmol/L、(134.74±12.65)μmol/L;VEGF的含量分别为(105.28±7.65) ng/L、(156.85±8.61) ng/L、(168.13±8.97) ng/L。Western blot检测结果显示,正常组、模型组和化疗组大鼠血管组织中TLR4蛋白相对表达水平分别为0.12±0.03、1.02±0.10、1.21±0.09;NF-κB p65蛋白相对表达水平分别为0.11±0.02、1.09±0.12、1.35±0.11。与正常组比较,模型组大鼠血浆中ET-1、NO、VEGF的含量和动脉组织中TLR4、NF-κB p65蛋白相对表达水平均增加;与模型组比较,化疗组大鼠血浆中ET-1、NO、VEGF含量和动脉组织中TLR4、NF-κB p65蛋白相对表达水平显著增加,以上指标差异均有统计学意义(P<0.05)。结论:化疗可加重大鼠肺癌模型血液高凝状态,使血管内皮功能紊乱,其机制可能与增加血管内皮ET-1、NO和VEGF含量,激活TLR4/NF-κB信号通路有关。

Objective:To observe the effect of chemotherapy on coagulation function and vascular endothelial function of rat lung cancer model,and analyze its mechanism of action.Methods:30 SD rats were randomly divided into normal group,model group and chemotherapy group,with 10 rats in each group.The model group and the chemotherapy group used the carcinogenic iodine oil liquid tracheal instillation method to establish a rat lung cancer model,while the normal group rats were only infused with iodine oil liquid.Rats in the chemotherapy group were injected with 5 mg/kg cyclophosphamide intraperitoneally,once a day,5 days a week for 8 consecutive weeks,and rats in the normal group and model group were injected with the same amount of normal saline.At two hours after the last administration,the plasma activated partial thromboplastin time(APTT),prothrombin time(PT),thrombin time(TT)and fibrinogen(FIB)were measured.ELISA method was used to detect the content of plasma endothelin(ET-1),nitric oxide(NO)and vascular endothelial growth factor(VEGF).Western blot was used to detect the expression levels of aortic endothelial Toll-like receptor 4(TLR4)and NF-κB p65.Results:Compared with the normal group,the APTT,PT,TT of the model group were shortened and FIB increased.Compared with the model group,the APTT,PT,TT of the chemotherapy group were further shortened,and the FIB increased.ELISA test results showed that the plasma ET-1 content of normal group,model group and chemotherapy group were(35.67±3.89)ng/L,(51.61±4.14)ng/L,(59.57±4.51)ng/L.NO content with(90.31±10.61)μmol/L,(124.92±13.26)μmol/L,(134.74±12.65)μmol/L.VEGF content were(105.28±7.65)ng/L,(156.85±8.61)ng/L,(168.13±8.97)ng/L.Western blot detection results showed that the relative expression levels of TLR4 protein in the vascular tissues of normal group,model group and chemotherapy group were 0.12±0.03,1.02±0.10,1.21±0.09.The relative expression levels of NF-κB p65 protein were 0.11±0.02,1.09±0.12,and 1.35±0.11.Compared with the normal group,the content of ET-1,NO,VEGF in the plasma and the relative expression level of TLR4,NF-κB p65 protein in the arterial tissue of the model group were increased.Compared with the model group,the content of ET-1,NO,VEGF in the plasma and the relative expression level of TLR4,NF-κB p65 protein in the arterial tissue of the chemotherapy group were significantly increased,and the differences of the above indexes were statistically significant(P<0.05).Conclusion:Chemotherapy can aggravate the blood hypercoagulability of rat lung cancer model and cause vascular endothelial dysfunction.The mechanism may be related to the increase of ET-1,NO and VEGF content in the vascular endothelium and activation of TLR4/NF-κB signaling pathway.