The Role of Factor V Leiden mutation In Recurrent and Spontaneous Abortion

Ahmad A. Al-Qudah

The Role of Factor V Leiden mutation In Recurrent and Spontaneous Abortion

- Identification of Factor V .- Identification of APCR and FVL .- Recurrent and Spontaneous Abortion .- Goal’s of our study

- Significance of work .- Patients and controls.- Samples .- Methodology.

- Budget - Bibliography .

Factor V

- Factor V (also called Labile Factor ) is a protein of the coagulation system.

- The gene is located on the first chromosome (1q23) and is homologous to coagulation factor VIII.

- consists of 25 exons, and the resulting protein has a relative molecular mass of approximately 330kDa.

- Factor V made primarily by cells in the liver , and circulates in plasma as a single-chain molecule with a plasma half-life of about 12 hours.

- able to bind to activated platelets , and activated by thrombin, degraded by activated protein C, one of the principal physiological inhibitors of coagulation .

Factor V

Activated protein C resistance (APCR)

- Activated protein C resistance (APCR) is an inherited hemostatic disorder characterized by the inability of Activated protein C to cleave Factor Va and/or Factor VIIIa.

- Allow for longer duration of thrombin generation and may lead to a hypercoagulable state, Resulting in an increased risk of venous thrombosis, which can cause problems with circulation.

- The disorder can be acquired or inherited, the hereditary form having an autosomal dominant inheritance pattern.

- Up to 64% of patients with venous thromboembolism might have activated protein C resistance.

Activated protein C resistance (APCR)

- Factor V Leiden is an inherited disorder of blood clotting.- Mutation of this gene is a single nucleotide polymorphism

(SNP).- As a missense substitution it changes a protein's amino acid

in which the Leiden variant of factor V cannot be inactivated and degraded by activated protein C.

- it is the most common hereditary hypercoagulability disorder.

Factor V Leiden

Recurrent and Spontaneous Abortion

- Thrombosis of placenta, umbilical cord, or the fetus. Can be a major cause of recurrent and spontaneous abortion.

- Activated protein C resistance is the most common genetic predisposition to thrombosis in European countries and the United States.

- Recurrent Abortion mean’s that there is a history of at least three Abortions, with no complete pregnancy .

- However, the association between the Factor V Leiden mutation and Abortion is still controversial.

Our study

1. Identification of Factor V and Protein C , and their Function in Coagulation system .

2. Identification of Coagulopathies and Factor V Leiden .

3. Identification of the Activated Protein C Resistance related to Factor V Leiden .

4. Investigate the Role of Factor V Leiden in Recurrent and Spontaneous Abortion .

Significance of work

- Many previous studies have shown that Activated Protein C Resistance , such as Activated Protein C Resistance related to Factor VIII mutation , is a disorder that leads to Abortion .

- Other studies have shown that Factor II mutation and other Coagulation Factors and Cofactors mutations , that lead to the Formation of Placental Thrombosis lead to Abortion .

- In this study , The Role of Activated Protein C Resistance related to Factor V Leiden mutation is going to be determined in Females with Recurrent and Spontaneous Abortion .

Significance of work

Patients and controls

- One hundred and twenty five Fertile women ( n = 125 ) .- Divided into : - Fifty ( 50 ) women with history of three or more Abortions - Fifty ( 50 ) with history of completed pregnancies and early Spontaneous Abortion .

- The control group consisted of twenty five ( 25 ) women with history of completed pregnancies without any problems or complications.

Samples

- One 5 ml EDTA tube sample will be Taken from each individual , for Factor V Leiden mutation Assay .

Methodology

Detection of Factor V Leiden mutation :

- Factor V Leiden mutation will be detected in patient's Blood using PCR and Cross Linking hybridization . - After DNA extraction , and beginning of Amplification , Oligonucleotides complementary to sequences within exon 10 and intron 10 of the factor V gene were synthesized. - PCR results loaded into wells Plate .

- Allele-specific capture probes complementary to nucleotides, either the normal factor V gene or the mutant factor V (Leiden) gene . - These probes contained a biotin molecule at the 3 terminus and ′a photoactive coumarin cross-linking agent that was positioned opposite the thymidine residue at certain position .

Methodology

- These are three types of Probes , Normal , mutant and Control set of both . - incubate in water bath for 30 min at 45° , then wash to remove unhybridized probes and any other unwanted materials .

Methodology

- binding of conjugate to the probe–target complex , wash to remove unbound conjugate, and development and detection of a fluorescent .

Methodology

1. Duga S, Asselta R, Tenchini ML. "Coagulation factor V". Int. J. Biochem. Cell Biol. (2005) 36 (8): 1393–9 .

2. Dahlbäck B. "The discovery of activated protein C resistance". J. Thrombosis. Haemost. (2003) 1 (1): 3–9

3. Bernadette F. Rodak, G.A.F., Elaine M. Keohane, Hematology: Clinical principles and applications. 3th ed. 2007: Elsevier Saunders.

4. Koster T, Rosendaal FR, De Ronde H, Briët E, Vandenbroucke JP, Bertina RM. "Venous thrombosis due to poor anticoagulant response to activated protein C: Leiden Thrombophilia Study". Lancet. 1993 Dec; 342 (8886–8887): 1503–6.

5. Nicolaes GA, Dahlbäck B. "Congenital and acquired activated protein C resistance". Semin Vasc Med. (2003) 3 (1): 33–46.

Bibliography