Identification of Factor V Leiden Genotypes in Normal Pools

Protein C serves as a natural anticoagulant, becoming active when thrombin binds to thrombomodulin on endothelial cell surfaces. Activated Protein C (APC) normally breaks down factor Va and factor VIIIa, inhibiting them reduce thrombin production and thus regulating anticoagulant activity. Conversely, poor APC cleavage can lead to decreased anticoagulant activity and increased blood coagulation, resulting in a hypercoagulable state known as “Resistance to APC (Rosen, SB, 1997 and Dahlback, B, 1999). APC resistance is a major risk factor for venous thromboembolism (VTE), including deep vein thrombosis (DVT) and pulmonary embolism (PE). Additionally, APC resistance can be inherited genetically( De Stefano, V and Leaone, G, 1995) or acquired through factors like oral contraceptives (OC) or hormonal therapy (HT) (Nicolaes,GA, 2003). One common cause of APC resistance is the Factor V Leiden (FVL) mutation, where Arg is replaced by Gly at position 506 (Factor V: Q506). The FVL mutation is inherited in an autosomal dominant manner. In this study, citrated plasma samples of 120 healthy individuals (median age: 42; age range: 24-62; number of female participants: 40 and male participants 80) were collected and tested for Prothrombin Time (PT), Activated Partial Thromboplastin Time (APTT), and Fibrinogen levels to assess blood clotting ability to prepare laboratory normal pool plasma. For all participants, PT (10.2-13.4 s), APTT (26-36 s), and Fibrinogen (2.5-4.4 g/L) were within normal ranges, indicating no abnormalities. However, medical chart review revealed that one participant (patient X) had a family history of DVT, OC/HT use, and the second participant (patient Y) had a history of DVT. Since PT and APTT clot times were normal, subset of 22 normal samples, including subject X and Y citrated plasma samples were further evaluated for coagulation clotting factors factor V and VIII levels and APTT-based APC Resistance test (APC-RV) for Factor V Leiden, Subjects X and Y coagulation clotting factors FVC levels were 78%, 96% & FVIIIC 95% & 112%, respectively and found to be in normal range of 50-150%.The subject X; patient Y; and 20 patients with no history of VTE, OC, or HT. APC ratios of 1.82 and 1.90 were obtained for subject X and Y, respectively, and these ratios fell within the laboratory-established FVL Heterozygous cutoff ratio of >1.4 to <=2.20. On the other hand, APC ratios of >=2.25 to <=4.2 were obtained for the no-history patients, indicating a FVL genotype. The two heterozygous phenotype patients, patients X and Y, were recommended for genetic testing to confirm the presence of the FVL mutation and removed from Normal Plasma Pool. In conclusion, this study demonstrates that it is important to evaluate personal and family medical history, to prepare a laboratory normal pool which is key factor to interpret clotting time values and if necessary, perform specialty screening tests.

No relevant conflicts of interest to declare.