Inactivation of the Coagulation Factor V (Essay Sample)

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Factor V
Introduction
The process of activation and inactivation of the coagulation factor V in the human blood is essential in terms of the regulation of blood. The main factor responsible for an activation of FV is Thrombin. This factor generates through a limited activated factor V molecule (FVa) known as proteolysis, which entails a light and heavy chain that Ca2+ ion holds together. The activated factor molecule makes use of its procaugulant function through playing the role of a non-enzymatic co-factor of Xa (FXa), serine protease factor. This is possible through the acceleration of the activation of prothrombin 103-105 folds. The activated protein, which associates with proteolysis in the 3-peptide bonds of FVa’s heavy chain, is responsible for catalysing the inactivation of FVa activity. The most fundamental reaction in the anticoagulant protein of C pathway is Proteolytic inactivation. Over the years, there have been reports regarding various human FV mutations. Among these mutations is the Factor V Cambridge (Arg306→Thr) a new factor that associates with the resistance of thrombosis and activated protein C.
Factor V structure
Steen et al reported that the I359T seemed to affect anticoagulation through two mechanisms. They impend the mediated down regulation APC of factor Va molecule and are also a poor APC cofactor for factor VIIIa down regulation. This explains I359T mutation association with thrombosis. Human beings FV gene coding located on chromosome 1 consist of approximately 80 kb spans and 25 exons. Transcription of human FV gene results to a mRNA of 6.5 kb. This predicts to 2224 amino acid proteins along with 28-amino acid leader peptide. This is a mature protein that has ~330 kD single chain polypeptide with a ~20 nmol/L plasma concentration mostly produced at the liver. The structure of FV is mosaic-like and similar to the structure of FVIII. It also has a domain structure where the FV and FVIII’s A and C domain share significant homology of approximately 40%.


Structural representation of FV and FVa
Factor v function
Activated FV (FVa) plays the role of FXa’s cofactor in the transformation of prothrombin to thrombin. The conversion takes place in the prothrombinase complex.
Factor v Cambridge treatment
According to Williamson et al, the characterization of factor v Cambridge as the reason for APC resistance proves the fact that the propensity of both arterial and venous thrombosis results from diverse genetic mutations. This mutation results to the change of arginine at position 306 to threonine, a change that results to the removal of the detection site for the limitation enzyme BstNI. Treatment of patients diagnosed with factor V Cambridge mutation resembles the treatment for FV: R506Q. Antithrombotic therapy is useful in acute thrombosis intervention followed by anticoagulation with warfarin. Some patients may undergo long-term secondary propholyxis using heparine or warfarine. The pharmacologic agent selected assists in monitoring.
Runge et al suggests that this form of mutation was only present in one of the patients from a group of 17 carefully selected patients, suffering from venous thrombosis along with a long-term resistance to APC in the absence of Gln506, which is a common mutation.1 In addition, factor V mutation was present in another individual, a first degree relative to the patient, who had APC resistance. In the process, various there was the exclusion of other factors that could result to APC resistance such as actor V HR2 haplotype and mutation that could occur at the Arg679 site. The mutation for factor V Cambridge was not present in 226 individuals who donated blood and in 585 venous thromboembolism patients following their screening. The first description of a mutation that affected the APC cleavage site for Factor V Cambridge involved Factor VThr306. In addition, it is the only mutation, apart from factor V Leiden, which associates with APC resistance. It is evident that this finding proves the physiologic importance the role of Arg306 APC-cleavage site when it comes to the prothrombinase complex. The finding is also essential as it supports the notion that a range of genetic mutations responsible for affecting fundamental sites in the cofactor of factor V can result to venous thrombosis and APC resistance.
In 1993, doctors reported a regular cause of familial thrombophilia, as the basis of poor anticoagulant response to APC. Activated Protein C resistance is present in approximately 3-5 percent of asymptomatic Caucasians. Castaman et al argue that it is also present in about 20% of various patients suffering from venous thrombosis. A single point mutation that occurs in the factor V gene causes approximately 95 % of APC resistance cases. 2 A change from G to A that takes place in exon 10 at the nucleotide 1691, results in the synthesis of factor V molecule variant known as Factor V Leiden, along with the substitution of Arg→Gln at position 506 of the amino acid. During the process of the prothrombinase complex assembly, factor V converts into an active Cofactor known as Va. The cleavage of cofactor Va through activated protein c at Arg506, followed by a cleavage that occurs at Arg306 and Arg679 limits the thrombin generation. The vitro experiments show that the cleavage at Arg506 does not affect any cofactor activity, but it is responsible for the exposure of inactivating cleavage sites particularly at Arg306. Both the potential of thrombin in Vitro and the danger that associates with thrombosis in Vivo increases due to the inactivation rate of factor VaGln506 and factor VaArg506. This is since VaGln506 activation rate is slower than the rate of factor VaArg506.
APC resistance in 5 -10 percent of patients characterised by deep vein thrombosis as well as those with APC resistance with no FVGln506 mutation could be due to various factors such as lupus anticoagulant activity, pregnancy or high level of factor VIII. The other cause of Activated protein C’s resistance, factor v protein, has been noted to be in patients with FVGln506 mutation along with HR2 haplotype. This is because these patients may experience lower ratios in terms of APC sensitivity but lack mutation, apart from FVGln506. In their study, Williamson et al carried out an experiment to identify mutations present in the sequence of the factor V gene comprising of the primary in activation site that occurs at Factor V Cambridge, in those patients that have cases of APC resistance and Venous thromboembolism. They selected patients from a large group, who had venous thromboembolism, based on definite APC resistance in the process where there was a lack of FVGln506 mutation.
In their study, Williamson et al measured the APC ratios of sencitivity in patients on samples of plasma from 602 patients successively investigated following a deep vein thrombosis diagnosis or pulmonary embolus diagnosis.3 During this time of blood sampling, the patients were not using any warfarin. Venography and ultrasound were responsible for determining deep vein thrombosis while ventilation-perfusion determined pulmonary embolus. According to their study, 424 patients had deep vein thrombosis while 178 had a diagnosis of symptomatic pulmonary embolism. They determined various aspects such as Standard APC sensitivity ratio, modified APC resistance assay, extended APC resistance assay and the activity of Plasma factor V coagulant. In addition, they determined natural anticoagulants, the FII20210 mutation and lupus anticoagulant activity, along with Factor VGln506, factor V exon 7 magnifications and analysis of V HR1/HR2 haplotype factor among others.10
Standard APC Sensitivity Ratio
There was a configuration of the samples from the patients twice at 2,500g during this process. This took place for approximately 10 minutes followed with freezing of platelet-poor plasma aliquots at ‒80°C until evaluation. Determining the APC sensitivity ratio occurred after filtering the plasma through a syringe filter of 0.2-μm with the aid of a Coated APC Resistance-C kit. The incubation of the plasma then followed with an equal ratio of partial thromboplastin reagent time for 5 minutes. The addition of CaCl2 initiated the clotting process. They achieved the clotting time using a ratio of time for clotting in the occurrence of APC divided by clotting time without APC. The resistance of APC significantly reduces due to the plasma filtration method. This results from the contamination of platelet filtration and increases the true APC resistance assay from 32% to 98%. Employing this method in a healthy control will result to an APC ratio that is greater by 2.2 than the result from unhealthy control.
Modified APC Resistance Assay
Dahlback et al assessed the resistance of APC in the occurrence of the factor V-depleted plasma with the aid of APC Resistance-C kit. They then pre diluted the plasma 1 in 5 using the factor V depleted plasma. This assisted in determining the sensitivity ratios in the standard assay. The results were that 40 of the selected patients showed a modified sensitivity ratio, which was less than 2.0. These patients had Gln506 mutation. The other unselected patients showed a sensitivity ratio that was greater than 2.2, and they had no Gln506 mutation.4
Extended APC resistance assay
Dahlback et al, describes an APC resistance assay, where there was incubation of platelet-poor plasma for 5 minutes using an equal amount of APTT reagent. Recording of the clotting time followed the addition of addition of CaCl2 enhanced with APC. This was over an ultimate range of concentration of 0-100 nmol/L.4
Family Study
Martinelli carried out an experiment on family study. The main patient, a 49-year-old man, had factor V Cambridge muta...