本文選題：達(dá)比加群 + 利伐沙班��； 參考：《北京協(xié)和醫(yī)學(xué)院》2016年博士論文

【摘要】：第一部分達(dá)比加群和利伐沙班對組織因子誘導(dǎo)的凝血酶生成和血小板聚集功能的影響目的：達(dá)比加群酯和利伐沙班是新近應(yīng)用于臨床的非維生素K依賴的直接凝血酶和直接因子Xa抑制劑。臨床研究已經(jīng)證明二者的療效不劣于甚至優(yōu)于傳統(tǒng)的抗凝藥物華法林。由于作用靶點的不同,二者抗凝效應(yīng)必然存在差異。本研究通過體外血漿凝血酶生成(TG)和全血血小板聚集率(PAgR)試驗評價達(dá)比加群和利伐沙班對組織因子誘導(dǎo)的凝血過程的影響,并探討凝血酶調(diào)節(jié)蛋白(TM)在抗凝藥物環(huán)境下對TG的調(diào)節(jié)作用。方法：枸櫞酸鈉抗凝管采集健康受試者外周靜脈全血,按以下三個方面處理：①分離乏血小板血漿(PPP),體外給予遞增濃度梯度(0,25,100,400,800 nmol/l)的達(dá)比加群和利伐沙班預(yù)處理,應(yīng)用校正的自動凝血酶生成圖(CAT)法檢測TG參數(shù),包括延遲時間(Lagtime)、內(nèi)源性凝血酶生成能力(ETP)、峰值(Peak)和達(dá)峰時間(Ttpeak);②枸櫞酸鈉抗凝全血在體外給予上述濃度梯度的達(dá)比加群和利伐沙班預(yù)處理,動態(tài)血小板計數(shù)法檢測全血PAgR;③在①基礎(chǔ)上加用等量的重組人凝血酶調(diào)節(jié)蛋白(rhs-TM,10 nmol/l),觀察TG變化。結(jié)果：①達(dá)比加群(0-800 nmol/l)濃度依賴性的延長血漿Lagtime和Ttpeak等時間參數(shù)。低濃度的達(dá)比加群(25-100 nmol/l)不降低血漿ETP(1737.6-1904.7 nmol-min)和Peak (337.7-364.6 nmol),高濃度的達(dá)比加群(100-800 nmol/L)濃度依賴性降低血漿ETP和Peak。利伐沙班(0-800nmol/l)延長Lagtime和Ttpeak及降低ETP和Peak均呈濃度依賴性的。以ETP計算,達(dá)比加群和利伐沙班的半數(shù)最大抑制濃度(IC50)分別為678.1±1.4 nmol/l和460.1±1.4 nmol/l,二者比值約1.47倍。②在空白對照組,組織因子可誘導(dǎo)78±5%的全血PAgR;達(dá)比加群和利伐沙班(0-800nmol/l)均可濃度依賴性的抑制組織因子誘導(dǎo)的PAgR, IC50分別為119.5±1.5和77.5±1.6nmol/l,二者比值約1.54倍。③rhs-TM不影響Lagtime和Ttpeak,但顯著降低ETP和Peak(分別下降22.4%和13.3%)。在rhs-TM環(huán)境下,低濃度的達(dá)比加群(25 nmol/l)有增加ETP(1568.97±117.99 vs 1478.65 ±73.77 nmol-min)和Peak(330.82±17.56 vs 299.46 ±6.35 nmol/l)的趨勢(P0.05)。不同的是,低濃度利伐沙班(25 nmol/)即可顯著降低ETP (748.42±368.97 vs 1478.65 ±73.77 nmol-min)和Peak(106.53±55.46 vs 299.46 ±6.35 nmol/l)(P均0.01)。以ETP計算,達(dá)比加群和利伐沙班的IC50分別為605.5±1.9 nmol/l和24.7±1.1 nmol/l,二者比值約24.5倍。結(jié)論：達(dá)比加群濃度依賴性的延遲TG的起始期,低濃度的達(dá)比加群(≤100nmol/l)對TG的擴(kuò)增期無影響。利伐沙班濃度依賴性的延遲TG的起始期、抑制TG擴(kuò)增期。利伐沙班和達(dá)比加群能濃度依賴性的抑制組織因子誘導(dǎo)的全血PAgR。利伐沙班對血漿TG和全血PAgR的抑制效應(yīng)強(qiáng)于達(dá)比加群。rhs-TM能增強(qiáng)利伐沙班的抗凝效應(yīng),對達(dá)比加群的抗凝效應(yīng)的影響較小。第二部分心房顫動患者應(yīng)用華法林起始抗凝治療過程中血漿內(nèi)源性凝血酶生成能力變化趨勢及臨床意義目的：具有高危血栓風(fēng)險的心房顫動患者積極抗凝治療可改善臨床預(yù)后。間接證據(jù)表明華法林起始應(yīng)用過程中可能出現(xiàn)反常高凝和增加血栓事件發(fā)生。本研究評價心房顫動患者應(yīng)用華法林起始抗凝階段血漿內(nèi)源性凝血酶生成能力(ETP)的變化趨勢,探討這一變化趨勢的臨床意義。方法：前瞻性收集具有抗凝適應(yīng)癥但近2周內(nèi)未行抗凝治療的心房顫動患者39例,按起始抗凝方案中是否聯(lián)合應(yīng)用低分子肝素(1OOAxaIU/kg)分成2組：A組：華法林組,26例；B組：聯(lián)合治療組,13例。A組按華法林給藥劑量分為三個亞組：A1：1.5mg qd,7例；A2:3mg qd,11例；A3:6mg qd×2d,然后3mg qd,8例。收集患者人口學(xué)特征、病史資料、基線血常規(guī)、D二聚體和纖維蛋白原水平。枸櫞酸鈉抗凝管采集患者基線和用藥后連續(xù)3天的靜脈全血。采用校正的自動凝血酶生成圖(CAT)法檢測ETP。同時間點檢測國際標(biāo)準(zhǔn)化比值(INR)、活化部分凝血活酶時間(APTT)、蛋白C(PC)和蛋白S(PS)等。多元線性回歸分析ETP與其它因素的關(guān)系。結(jié)果：除了糖尿病病史兩組之間存在顯著差異外(3.8% vs 30.8%,P=0.035),其它基線特征、基線INR、APTT、PC、PS和ETP等均未見顯著差異。2組患者INR動態(tài)比較無顯著差異(累計增幅33.8%vs 34.8%, P=0.947); APTT在華法林組表現(xiàn)為逐步升高的過程；在聯(lián)合治療組第1天即顯著升高(增幅14.5%vs 3%,P=0.026)并維持這一水平,至第3天兩組APTT無顯著差異(增幅13.5%vs 15.84%,P=0.628)；兩組PC和PS活性均呈現(xiàn)緩慢下降趨勢,PC/PS累計降幅均無顯著差異(分別為-28.6%vs-23.9%,-18.6%vs-14.5%：P值分別為0.505和0.565),其中第2天華法林組PC日降幅顯著高于聯(lián)合治療組(-14.7%vs -7.0%,P=0.033)。聯(lián)合治療組ETP隨時間推移直線下降；華法林組ETP的降幅在第2天呈小幅波動,兩組患者3天ETP累計降幅無顯著差異(-30.5% vs -36.8%,P=0.273)。多元線性回歸分析提示基線ETP與紅細(xì)胞(RBC)計數(shù)(p=257.6,P=0.001)和INR獨立相關(guān)(p=-1425.8,P=0.001)；第1天和第2天ETP的日降幅與同時段INR變化幅度獨立負(fù)相關(guān)(分別β=-0.946,-0.231；P=0.002,0.004),3天累計ETP降幅與同時段PC降幅獨立負(fù)相關(guān)(p=-0.378,P=0.002)。亞組分析發(fā)現(xiàn),華法林對PC、PS和ETP的抑制均呈現(xiàn)劑量依賴性。A1組3天ETP累計降幅顯著低于A2和A3組(分別為-20.4%,-34.1%和-34.3%)；A1和A2組ETP在第3天出現(xiàn)最大降幅(分別為-8.7%和-21.7%)；A3組ETP在第1天的降幅最大(-17.5%)。結(jié)論：小劑量的華法林起始抗凝治療過程中ETP最大降幅出現(xiàn)在第3天,大劑量的華法林在第1天即可獲得較顯著的ETP降幅。華法林對ETP、PC和PS活性的影響均呈現(xiàn)劑量依賴性。心房顫動患者應(yīng)用華法林起始抗凝過程中ETP的日降幅呈現(xiàn)小幅波動,該現(xiàn)象可能通過聯(lián)合低分子肝素治療獲得改善。RBC計數(shù)和INR值影響基線ETP水平；而華法林起始給藥過程中INR值和PC活性的變化與ETP的變化相關(guān)。
[Abstract]:The first part of the effect of dabiga group and LEV Shaaban on tissue factor induced thrombin formation and platelet aggregation purpose: dabig and LEV Shaaban are newly used direct thrombin and direct factor Xa inhibitors for clinical non vitamin K dependence. Clinical studies have shown that the efficacy of the two is not inferior to even superior. The anticoagulant effect of the traditional anticoagulant drug, warfarin, is necessarily different from the two anticoagulant effects. In this study, the effects of dabiga group and LEV Shaaban on the coagulation process induced by tissue factors were evaluated by plasma thrombin formation (TG) and blood platelet aggregation (PAgR) test in vitro, and the thrombin regulatory protein (TM) was discussed. The regulation of TG under the anticoagulant environment. Methods: the sodium citrate anticoagulant tube was used to collect the peripheral venous blood of the healthy subjects. According to the following three aspects: (1) the isolated platelet plasma (PPP) was separated, and the dabiga group and LEV Shaaban pretreated with the increasing concentration gradient (0,25100400800 nmol/l) were pretreated in vitro, and the corrected automatic coagulation was applied. TG parameters were detected by CAT, including delayed time (Lagtime), endogenous thrombin generation capacity (ETP), peak (Peak) and peak time (Ttpeak); (2) sodium citrate anticoagulant whole blood was given the above concentration gradient of dabiga group and ribeasta preconditioning in vitro, dynamic platelet counting method was used to detect PAgR in whole blood; (3) on the basis of 1 The same amount of recombinant human thrombin regulated protein (rhs-TM, 10 nmol/l) was added to observe the changes in TG. Results: (1) dabiga group (0-800 nmol/l) concentration dependent prolongation of plasma Lagtime and Ttpeak time parameters. Low concentration dabia group (25-100 nmol/l) did not reduce plasma ETP (1737.6-1904.7 nmol-min) and Peak (337.7-364.6), high concentration Dabiga group (100-800 nmol/L) concentration dependent reduced plasma ETP and Peak. Lev Shaaban (0-800nmol/l) prolonged Lagtime and Ttpeak and reduced ETP and Peak in a concentration dependent manner. In ETP, the maximum inhibitory concentration of dabiga group and LEV Shaaban (IC50) is 678.1 + 1.4 nmol/l and 460.1 + 1.4 nmol/l, and the two ratio is about 1.47 times. (2) in the blank control group, the tissue factor could induce 78 + 5% of the whole blood PAgR; dabiga group and LEV Shaaban (0-800nmol/l) could inhibit the concentration dependent tissue factor induced PAgR, IC50 was 119.5 + 1.5 and 77.5 + 1.6nmol/l respectively, the ratio of two was about 1.54 times. (3) rhs-TM did not sound Lagtime and Ttpeak, but significantly decreased ETP and Peak (respectively decreased. 22.4% and 13.3%). Under the rhs-TM environment, the low concentration dabiga group (25 nmol/l) has a trend of increasing ETP (1568.97 + 117.99 vs 1478.65 + 73.77 nmol-min) and Peak (330.82 + 17.56 vs 299.46 + 6.35 nmol/l). And Peak (106.53 + 55.46 vs 299.46 + 6.35 nmol/l) (P 0.01). The IC50 of dabiga group and LEV Shaaban's IC50 were 605.5 + 1.9 nmol/l and 24.7 + 1.1 nmol/l respectively. The ratio of two was about 24.5 times respectively. Conclusion: dabiga group concentration dependent delayed TG initiation period and low concentration Dabi group (< < < 100nmol/l) have no effect on TG amplification period. The starting period of Lev Shaaban concentration dependent delayed TG inhibited the TG amplification period. The inhibitory effect of Lev Shaaban and dabiga group on the inhibitory effect of PAgR. Lev Shaaban on plasma TG and whole blood PAgR was stronger than that of dabiga group.Rhs-TM to enhance the anticoagulant effect of Lev Shaaban, and the anticoagulant effect on dabiga group The change trend and clinical significance of plasma endogenous thrombin formation in second parts of atrial fibrillation patients during the initial anticoagulant treatment of atrial fibrillation: positive anticoagulant therapy with high risk of thrombus risk in patients with atrial fibrillation can improve clinical prognosis. Abnormal hypercoagulability and increased thrombotic events occurred. This study evaluated the changes in plasma endogenous thrombin formation (ETP) in warfarin initial anticoagulant stage in patients with atrial fibrillation, and explored the clinical significance of this trend. Method: prospective collection of atrial fibrillation with anticoagulant therapy in the near 2 weeks without anticoagulant therapy 39 patients were divided into 2 groups according to the combined application of low molecular weight heparin (1OOAxaIU/kg) in the initial anticoagulant regimen: group A: warfarin group, 26 cases; group B: combined treatment group; 13 cases of group.A were divided into three subgroups according to the dosage of warfarin: A1:1.5mg QD, 7 cases; A2:3mg QD, 11 cases; A3:6mg QD x 2D, then 3mg 8. Signs, medical history data, baseline blood routine, D two polymer and fibrinogen level. The sodium citrate anticoagulant tube collected the patient's baseline and 3 days of continuous venous blood after 3 days of medication. The revised automatic thrombin generation chart (CAT) was used to detect the international normalized ratio (APTT), the activated partial thromboplastin time (APTT), and the protein C (PC) at the same time point. Protein S (PS) and so on. Multivariate linear regression analysis of the relationship between ETP and other factors. Results: there were significant differences between the two groups (3.8% vs 30.8%, P=0.035), other baseline features, baseline INR, APTT, PC, PS and ETP, and no significant difference in INR dynamics (cumulative increase 34.8%). APTT in the warfarin group showed a gradual increase in the performance of the warfarin group, which was significantly increased (14.5%vs 3%, P=0.026) and maintained this level in the first days of the combined treatment group, and there was no significant difference between the two groups (13.5%vs 15.84%, P=0.628) in the third day group, and the two groups of PC and PS activity showed a slow decline trend, and there was no significant difference in the cumulative decrease of PC/PS. The value of -18.6%vs-14.5%:P was 0.505 and 0.565 respectively, and the decrease of PC day in second days was significantly higher than that in the combined treatment group (-14.7%vs -7.0%, P=0.033). The ETP in the combined treatment group decreased linearly with time, and the decrease of ETP in the warfarin group was slightly fluctuating in second days, and there was no significant difference between the two groups of patients at 3 days. -30.5% vs -36.8%, P=0.273). Multivariate linear regression analysis suggested that baseline ETP and RBC count (RBC) count (p=257.6, P=0.001) and INR independent correlation (p=-1425.8, P=0.001); first days and second days, the daily decline of ETP is independent of the amplitude of simultaneous variation (beta =, respectively), and 3 days' cumulative decline and simultaneous decline Independent negative correlation (p=-0.378, P=0.002). The subgroup analysis found that Hua Falin's inhibitory effect on PC, PS and ETP was significantly lower in the dose dependent.A1 group than the A2 and A3 groups at 3 days (-20.4%, -34.1% and -34.3%). 5%). Conclusion: the maximum ETP decline occurred during the small dose of Hua Falin initial anticoagulant therapy in third days, the large dose of Hua Falin could obtain a significant ETP decline in first days. The effect of Hua Falin on the activity of ETP, PC and PS showed a dose dependence. The decrease of the daily decrease of ETP during the application of the initial anticoagulant process in the patients with atrial fibrillation was small. Amplitude fluctuation, this phenomenon may improve the.RBC count and INR value by combined low molecular weight heparin therapy, and the changes of INR and PC activity during the initial administration of warfarin are related to the changes of ETP.

【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：R541.75

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