Interaction between warfarin and nonsteroidal anti-inflammatory drugs (NSAIDs) in rats

In fed rats, the following NSAIDs were administered orally 24 hr before or 18 hr after the intraperitoneal administration of 1.34 mg/kg warfarin: phenylbutazone, 150 mg/kg; diflunisal, 75 mg/kg; ibuprofen, 150 mg/kg; acetylsalicylic acid, 300 mg/kg; indomethacin, 8 mg/kg; tolmetin sodium, 50 mg/kg; ketoprofen, 8 mg/kg; and amfenac sodium, 8 mg/kg. The elevation of the 24-hr prothrombin time was indicative of the effect of the warfarin. Warfarin-treated fasted rats showed a significantly higher prothrombin time than warfarin-treated fed rats. Interaction with phenylbutazone and warfarin occurred in fed and not in fasted rats when administered 18 hr after administration of the warfarin. At the 24-hr pretreatment time, only phenylbutazone significantly reduced the elevated prothrombin time. With the exception of amfenac sodium, all the NSAIDs significantly enhanced the elevated prothrombin time when administered 18 hr after warfarin. Their decreasing order of activity in enhancing the elevated prothrombin time was phenylbutazone, diflunisal, acetylsalicylic acid, ibuprofen, indomethacin, tolmetin sodium, and ketoprofen. The results indicate that the rat is more sensitive than the human to the interaction between warfarin and NSAIDs.     

The submicrosomal site for the conversion of prothrombin precursor to biologically active prothrombin in rat liver

The submicrosomal site for the conversion of prothrombin precursor to prothrombin in rat lever has been investigated by subcellular fractionation techniques.Prothrombin precursor activity could be detected in the luminal as well as the membrane fraction of the rough microsomes. The corresponding fractions from smooth microsomes did not exhibit any activity. After warfarin treatment of the rats, the concentration of prothrombin precursor in rough microsomes increased rapidly from approx. 2 to 6–8 h, when a plateau was reached. In smooth microsomes, prothrombin precursor activity appeared 1 h after injection of warfarin, and increased to a plateau reached after about 4 h. The total activity of prothrombin precursor at the plateau obtained after warfarin treatment was 4–5 times higher in the rough luminal fraction than in the corresponding smooth fraction.The vitamin K-dependent carboxylase activity was localized to the rough microsomes. The enzyme system was associated with the membrane, mainly at the luminal side, whereas the substrate appeared to be localized in both the luminal and membrane fraction.The results indicate that the conversion of prothrombin precursor to biologically active prothrombin occurs at a late stage in the rough endoplasmic reticulum or at a transition between rough and smooth endoplasmic reticulum.     

Ineffectiveness of phylloquinone epoxide as an inhibitor of prothrombin synthesis in the rat

Phylloquinone epoxide (vitamin K₁-oxide), a metabolite of phylloquinone, does not inhibit prothrombin synthesis when administered in high doses to Sprague-Dawley and warfarin-resistant rats. Further, it does not accumulate to presumed inhibitory levels in the livers of rats given physiological doses of ³H-phylloquinone when they are anticoagulated with warfarin. These data do not support the Bell-Matschiner hypothesis that warfarin exerts its action by inhibiting the vitamin K oxide reductase which results in the accumulation of vitamin K oxide and the inhibition of vitamin K at its active site. Rather, our data support the view that vitamin K and warfarin combine at different sites with a single regulatory protein which serves as a conformational switch for prothrombin synthesis.     

Inhibition by warfarin of prothrombin synthesis and of lipid-saccharide synthesis in rat liver

In vivo and in vitro studies using [³H]glucosamine incorporation into prothrombin and into glycolipids were conducted in rat liver to determine the role of lipid-saccharides in the biosynthesis of prothrombin.In vivo studies demonstrated that 10 mg warfarin/kg inhibited the incorporation of radiolabeled glucosamine into liver prothrombin and glycolipids. This inhibition was similar to the kinetics of inhibition of prothrombin synthesis in the liver.In vitro studies demonstrated a time-dependent increase in the incorporation of radiolabeled glucosamine into lipid-saccharides and prothrombin. This incorporation was inhibited 50% by 5 · 10^?4 M warfarin. Warfarin also inhibited the incorporation of radiolabeled glucosamine into glycolipids in a dose-related manner.In all studies, vitamin K-1 reversed the inhibition of glucosamine incorporation into glycolipids and into prothrombin.     

Association of CYP2C9 gene polymorphism with bleeding as a complication of warfarin therapy

The aim of this study was to determine the association of bleeding as a complication of warfarin therapy with polymorphism of CYP2C9 gene (alleles 1, 2 and 3). The CYP2C9 is the main enzyme for warfarin metabolism. Study included 181 patients receiving warfarin for at least one month. Allele 1 of CYP2C9 gene (in 94.5%) and genotype *1/*1 (57.5%) prevailed. Allele 3 was found in 12.7% patients. Bleeding side-effects occurred in 18 patients (10%). Patients with allele *1 needed significantly higher maintenance warfarin dose (p=0.011). Those with allele *3 had significantly lower maintenance warfarin dose (p=0.005) and higher prothrombin time (PT) at induction (p=0.034). Bleeding occurred significantly more often in those with lower maintenance warfarin dose (p=0.017). Patients with allele *3 had increased risk of bleeding, with marginal significance (p=0.05). Polymorphism of CYP2C9 could determine dose of warfarin therapy and thus it could be related to the risk of bleeding complications. Allele *3 carriers need lower warfarin dose. Therefore, initially reduced warfarin induction dose in allele *3 carriers could avoid more prolonged PT and decrease the risk of bleeding complication.     

Flexible induction dose regimen for warfarin and prediction of maintenance dose.

Fifty patients with venous thromboembolic disease being treated by heparin infusion received a three day warfarin induction regimen tailored according to the prothrombin time (British comparative ratio) measured on days 2 and 3. A prediction of the final maintenance dose of warfarin was made on the basis of a prothrombin time measured on day 4. All patients were safely anticoagulated by day 6, and the prediction was accurate to within 1 mg in 46 patients. Predicted and actual maintenance doses were closely related (r = 0.867; n = 50; p less than 0.001). This scheme should prove helpful in the control of anticoagulation, particularly in patients likely to be sensitive to warfarin, and should shorten hospital stay.     

A Comparison of Warfarin Sodium and Bishydroxycoumarin in Long-Term Anticoagulant Therapy

Warfarin sodium was compared with bishydroxycoumarin (Dicumarol) in 16 patients on long-term anticoagulant therapy. When the patients were changed from bishydroxycoumarin to warfarin sodium there was no improvement in control of their prothrombin times. It was found that 5 mg. of warfarin had slightly less effect than 50 mg. of bishydroxycoumarin. It was concluded that the drugs were equally effective in long-term anticoagulant therapy. The metabolism of the ingested drug was more important than absorption in determining the control of the patients'' prothrombin times.     

Optimal loading dose for the initiation of warfarin: a systematic review

Background

Selection of the right warfarin dose at the outset of treatment is not straightforward, and current evidence is lacking to determine the optimal strategy for initiation of therapy.

Methods

We included randomized controlled trials in patients commencing anticoagulation with warfarin, comparing different loading dose or different regimens. We searched Medline, EMBASE, the Cochrane Library and the NHS Health Economics Database up to June 2009. Primary outcomes were time to stable INR and adverse events. We summarised results as proportion of INRs in range from date of initiation and compared dichotomous outcomes using relative risks (RR) and calculated 95% confidence intervals (CIs).

Results

We included 11 studies of 1,340 patients newly initiated on warfarin. In two studies that used single INR measures, a loading dose of 10 mg compared to 5 mg led to more patients in range on day five. However, in two studies which measured two consecutive INRs, a loading dose of 10 mg compared to 5 mg did not lead to more patients in range on day five (RR = 0.86, 95% CI, 0.62 to 1.19, p = 0.37). Patients receiving a 2.5 mg initiation does took longer to achieve the therapeutic range, whilst those receiving a calculated initiation dose achieved target range 0.8 days quicker (4.2 days vs. 5 days, p = 0.007). More elderly patients receiving an age adjusted dose achieved a stable INR compared to the Fennerty protocol (48% vs. 22% p = 0.02) and significantly fewer patients on the age adjusted regimens had high out-of-range INRs. Two studies report no significant differences between genotype guided and 5 mg or 10 mg initiation doses and in the one significant genotype study the control group INRs were significantly lower than expected.

Conclusion

Our review findings suggest there is still considerable uncertainty between a 10 mg and a 5 mg loading dose for initiation of warfarin. In the elderly, lower initiation doses or age adjusted doses are more appropriate, leading to less higher INRs. Currently there is insufficient evidence to warrant genotype guided initiation, and adequately powered trials to detect effects on adverse events are currently warranted.     

Anticoagulothérapie orale: analyse de l'efficacité d'un guide de posologie.

An empirical protocol for dosage adjustment in warfarin therapy was evaluated by means of retrospective analysis of the medical records of 106 outpatients at an anticoagulation clinic followed from January 1981 to August 1984. There were 203 cases in which the prothrombin time was abnormal (below the therapeutic range in 123 cases and above the range in 80). The protocol was successful in about 60% of cases in which the prothrombin time before dosage adjustment was below the therapeutic range and in 80% of cases in which it was above the range. Statistical analysis showed that the success rate could be improved by more precise definition of the percentage of dosage change in certain situations. However, individual variation in response of the prothrombin time to a standard dosage change limits the success that can be expected after the initial adjustment. We propose an improved protocol for adjustment of warfarin dosage.     

肝癌大鼠异常凝血酶原的研究

用灵敏的凝血酶原蛇毒激活测定法,观察二乙基亚硝胺诱发大鼠肝癌过程中,血浆异常凝血酶原及肝组织内凝血酶原前体含量的变化。血浆异常凝血酶原从诱癌第4周开始持续上升至第13周;第15周有所下降,第20周又开始回升。而在此过程中,肝组织内凝血酶原前体含量变化不明显,而在诱癌20周的肝癌结节内,凝血酶原前体显著堆积。同时,还研究了肝再生过程中大鼠血桨异常凝血酶原及肝内凝血酶原前体的变化。华法令处理的大鼠作为阳性对照。     

The effect of warfarin and vitamin K-1 on the carboxylation and glycosylation of prothrombin in vivo

The effect of warfarin and vitamin K-1 on the carboxylation and glycosylation of prothrombin in the rat in vivo was investigated. Neither warfarin nor vitamin K-1 has an effect on carboxylation. However, warfarin inhibited glycosylation 80–90% and this inhibition was readily reversed by the administration of vitamin K-1.     

Growth inhibitory actions of prothrombin on normal hepatocytes: influence of matrix

Most hepatomas have a defect in prothrombin carboxylation, and can secrete under-carboxylated prothrombin or des-gamma-carboxy-prothrombin (DCP), the function of which is unknown. We considered that the prothrombin-DCP axis might also be involved in growth control. Hepatocytes and hepatoma cells were treated with prothrombin and DNA synthesis and cytoskeletal changes were studied. Prothrombin inhibited DNA synthesis in hepatocytes on fibronectin, but not collagen matrix. Hepatoma cell lines were not inhibited. We found that hepatoma cell matrix conferred resistance to hepatocytes. Prothrombin decreased fibronectin but not collagen amounts, but only in the presence of hepatocytes and not hepatoma cells, indicating that it has a differential action on matrix proteins. It also caused changes in cell shape and actin depolymerization. In vivo, there was a decrease in plasma prothrombin activity after a partial hepatectomy (PH), concomitant with the peak of DNA synthesis in the hepatocytes at 24h after PH. Injection of warfarin at the time of PH, further inhibited PT activity and enhanced this 24h peak of DNA synthesis. Furthermore, repeated injection of prothrombin lowered the peak DNA synthesis after PH. The data support the hypothesis that prothrombin can act as a hepatocyte growth inhibitor, likely at the level of fibronectin loss and result in cytoskeletal changes. Hepatomas resist this action, possibly due to their different matrix proteins. This represents a novel mechanism for growth regulation and provides a possible biological significance for the tumor marker DCP.     

Prothrombin biosynthesis: characterization of processing events in rat liver microsomes

Plasma and hepatic microsomal forms of rat prothrombin have been compared by sodium dodecyl sulfate-polyacrylamide electrophoresis and isoelectric focusing. The major prothrombin species that accumulated in the microsomes of rats treated with warfarin had a molecular weight of 78 500 and a pI in 8 M urea of 6.3-6.5. Plasma prothrombin had a molecular weight of 83 500 and a pI of 5.3-5.7. Microsomes from normal rat liver contain a second pool of precursor with a molecular weight of 83 500, and digestion with the glycosidase Endo H indicated that this form has been processed to contain complex carbohydrates, while the Mr 78 500 form is a high mannose form and is the substrate for the vitamin K dependent carboxylase. Treatment of rats with tunicamycin revealed that glycosylation was not essential for carboxylation or secretion from the liver. Comparison of the aglyco forms of prothrombin and its precursors suggests that the intracellular forms contain a basic, Mr approximately 1500 peptide that is missing from the plasma form of prothrombin.     

Association of congenital deficiency of multiple vitamin K-dependent coagulation factors and the phenotype of the warfarin embryopathy: clues to the mechanism of teratogenicity of coumarin derivatives.

We have evaluated a boy who had excessive bleeding and bruising from birth and showed markedly prolonged prothrombin times, partially correctable by oral vitamin K administration. Additional laboratory studies demonstrated decreased activities of plasma factors II, VII, IX, and X; near normal levels of immunologically detected and calcium binding-independent prothrombin; undercarboxylation of prothrombin; excess circulating vitamin K epoxide; decreased excretion of carboxylated glutamic acid residues; and abnormal circulating osteocalcin. These results all are consistent with effects resulting from decreased posttranslational carboxylation secondary to an inborn deficiency of vitamin K epoxide reductase. This individual also had nasal hypoplasia, distal digital hypoplasia, and epiphyseal stippling on infant radiographs, all of which are virtually identical to features seen secondary to first-trimester exposure to coumarin derivatives. Therefore, by inference, the warfarin embryopathy is probably secondary to warfarin's primary pharmacologic effect (interference with vitamin K-dependent posttranslational carboxylation of glutamyl residues of various proteins) and may result from undercarboxylation of osteocalcin or other vitamin K-dependent bone proteins.     

Canadian atrial fibrillation anticoagulation study: were the patients subsequently treated with warfarin? Canadian Atrial Fibrillation Anticoagulation Study Group.

OBJECTIVE: To determine the effect of the results of clinical trials on the behaviour of patients and physicians, the authors ascertained the proportion of patients participating in the Canadian Atrial Fibrillation Anticoagulation (CAFA) study who started or continued warfarin therapy at the end of the study and identified factors affecting the decision to use or not use warfarin. The CAFA study was a double-blind, randomized, placebo-controlled, multicentre study to evaluate the efficacy of warfarin in preventing stroke among patients with nonrheumatic atrial fibrillation. Recruitment and follow-up were stopped early because two other similar studies had shown a decrease in the rate of stroke among patients treated with warfarin. DESIGN: Mail survey 21 months after the end of the study. PARTICIPANTS: The personal physicians of 336 patients who had participated in the CAFA study. OUTCOME MEASURES: Type of antithrombotic therapy the patients had received since the CAFA study ended for patients who were not receiving warfarin, the reasons they were not. RESULTS: Questionnaires concerning 254 (76%) of the patients who had participated in the study were returned. Since the end of the CAFA study, 153 (60%) of these patients had been treated continually with warfarin, 14 (6%) had been treated with warfarin but had subsequently stopped taking it, 59 (23%) had taken acetylsalicylic acid (ASA) continually, 5 (2%) had been taking ASA but had subsequently stopped taking it, and 23 (9%) had not taken either drug. The responding physicians stated that 58 (67%) of the patients who were not treated with warfarin did not wish to take the drug. The patients who had received warfarin during the CAFA trial were more likely to be treated with warfarin after the trial (75%) than were those who had received a placebo (56%) (p = 0.001). The probability of the patients'' being treated with warfarin also depended on which study centre they had been treated in (p = 0.001). CONCLUSIONS: Of the patients in the CAFA study for whom questionnaires were received, only 167 (66%) had been treated with warfarin after the end of the study. The patients were more likely to have been treated with warfarin after the study if they had received warfarin during the study. The positive results of clinical trials, on their own, are not enough to fully change the behaviour of patients and physicians.     

Procoagulant effect of phenobarbital in rats

Daily administration of phenobarbital, 75 mg/kg ip for 4 days, to adult male Sprague-Dawley rats produced a pronounced increase of prothrombin complex activity (PCA) in plasma, i.e. a decrease of the prothrombin time. This effect persisted for 4 to 5 days after the last dose of phenobarbital. The rate constant for the decline of PCA after administration of a PCA synthesis-blocking dose of warfarin was not affected by treatment with phenobarbital but the rate of synthesis of PCA was increased appreciably. Thus, the phenobarbital-induced increase of PCA was caused by increased synthesis of one or more clotting factors.     

Ingestion of superwarfarin leading to coagulopathy: a case report and review of the literature

Superwarfarins are found in many pesticides, including D-con, Prufe I and II, Ramik, Talon-G, Ratak, and Contrac. Ingestion of can lead to significant morbidity and even mortality. Physicians need to consider this diagnosis in any patient presenting with coagulopathy of unclear etiology. We present a patient with superwarfarin-induced coagulopathy and review previous cases in adults in the literature. The patient is a 60-year-old man who presented to our medical center with painless hematuria. Laboratory studies revealed an elevated prothrombin time (PT) (42.5 seconds), partial thromboplastin time (PTT) (64.6 seconds), and international normalized ratio (INR) of 7. Liver-associated enzymes were normal, and complete blood cell count (CBC) showed no evidence of disseminated intravascular coagulation. Subsequent work-up included the absence of an inhibitor by mixing study and deficiencies of vitamin K-dependent coagulation factors. The patient's warfarin level was negative. A brodifacoum level was positive, confirming superwarfarin-induced coagulopathy. The patient is currently doing well with normal coagulation studies after receiving high doses of vitamin K for several weeks. The cause of his exposure to superwarfarin remains uncertain. Physicians need to be cognizant of this unusual cause of coagulopathy in adults. The appropriate diagnostic work-up and unique features of therapy are discussed.     

Exposure of the pregnant rat to warfarin and vitamin K1: an animal model of intraventricular hemorrhage in the fetus

Pregnant Sprague-Dawley rats were given daily oral doses of sodium warfarin (100 mg/kg) and concurrent intramuscular injections of vitamin K1 (10 mg/kg). This dosing regimen did not have any apparent deleterious effect on the dams and did not affect the fetuses when administered from day 1 to day 12 of pregnancy. However, similar treatment from day 9 to 20 caused hemorrhage in the fetuses examined on day 21 of gestation. There were no hemorrhages in the control fetuses from dams receiving vitamin K1 only. The lowest effective dose of warfarin, in conjunction with daily doses of vitamin K1, was 3 mg/kg. This dose caused hemorrhage in 28% of fetuses; the incidence of affected fetuses was not further increased by doses of warfarin up to 100 mg/kg. Hemorrhages affected the fetal brain, face, eyes, and ear and occasionally the limbs. Brain hemorrhages were frequently intraventricular and caused various degrees of hydrocephaly. Bony defects were not a feature of prenatal exposure to warfarin. These results show that prenatal exposure of the rat to warfarin and vitamin K duplicates the hemorrhagic abnormalities and pathology associated with prenatal exposure to warfarin in the human. It did not induce bony or facial defects probably because the vitamin K-dependent components of bone development occur postnatally in the rat. This model should allow detailed determination of the role of vitamin K-dependent proteins in development.     

Purification and characterization of a prothrombin activator from the venom of the Australian brown snake, Pseudonaja textilis textilis

A simple procedure, involving chromatography on concanavalin A-Sepharose and gel filtration, has been developed for the purification of a prothrombin activator from the venom of the Australian brown snake Pseudonaja textilis textilis. The prothrombin activator, which is a major venom component, is a high molecular weight protein (Mr greater than or equal to 200,000) which yields a number of subunits when examined by SDS-PAGE. It is related antigenically to the venom prothrombin activator of the taipan Oxyuranus scutellatus. P. textilis prothrombin activator is able to coagulate citrated plasma, warfarin plasma, and Factor V- and Factor X-deficient plasmas; to convert purified human prothrombin to thrombin; and to hydrolyse the peptide p-nitroanilide substrate S-2222. Calcium ions and phospholipids had little if any effect on the rates of coagulation of citrated plasma or S-2222 hydrolysis catalysed by this enzyme.     

利伐沙班与华法林对高龄非瓣膜性房颤患者D-二聚体、NT-proBNP水平的影响

目的:探讨利伐沙班与华法林对高龄非瓣膜性房颤(NVAF)患者血浆D-二聚体(D-D)、N末端B型利钠肽原(NT-pro BNP)水平的影响及其临床疗效。方法:选取我院2015年1月~2016年11月收治的146例高龄NVAF患者,采取随机数字表法均分为两组。华法林组予以华法林抗栓治疗,利伐沙班组采取利伐沙班抗栓治疗。记录比较两组治疗期间栓塞、出血情况及不良反应,以及治疗前后血浆D-二聚体(D-D)、N末端B型利钠肽原(NT-pro BNP)水平的变化情况。结果:两组治疗后栓塞发生率比较,差异无统计学意义(P0.05)。利伐沙班组出血发生率(4.1%)低于华法林组(15.1%),差异具有统计学意义(P0.05)。与治疗前相比,两组治疗后血浆D-D和NT-pro BNP水平均降低,差异具有统计学意义(P0.01);治疗后,两组血浆D-D和NT-pro BNP水平比较,差异无统计学意义(P0.05)。两组不良反应发生率对比,差异无统计学意义(P0.05)。结论:与华法林相比,高龄非瓣膜性房颤应用利伐沙班抗栓治疗在患者耐受性与预防血栓栓塞方面优势相当,但利伐沙班更能有效降低患者出血风险。