33.3 Animal model for haemophilia

33.3.1 Development of Factor IX deficient mice

A factor IX-deficient mouse model for hemophilia B gene therapy was developed by Wang et al., in the year 1997 to study the bleeding disorder (hemophilia B). The factor IX gene was knocked out and this knock-out mice (−/−) was then comparatively studied with the control wild type (+/+) and heterozygous mice (+/−).

The mice were traumatized by injuring their feet using a mouse restrainer. In case of knock-out mice swollen extremities and hematoma were very prominent whereas, these were found to be reduced in the wild type and heterozygous mice. Using Activated Partial Thromboplastin Time (APTT) assay the factor IX coagulant activity in all three mice wild type (+/+), heterozygous (+/−) and knock-out (−/−) was quantitatively estimated and was found to be 92.1 ± 19.1% for the wild type (+/+), 52.9 ± 10.4% for the heterozygous (+/−) and 5.4 ± 2.9% for the knock-out (−/−) mice.

This study was successful in terms of creating Factor IX knock-out mice.

33.3.2 Development of WAG-F8 (m1Ycb) rats harboring a factor VIII gene mutation

In 2010 Booth et al. , developed a rat model for factor VIII in order to study the hemophilia A condition. The main objective of this study was

• •  The determination of precisely specific factors in the coagulation mechanism in the WAG/RijYcb rats.

•  Finding out the genetic defect associated or responsible for the hereditary clotting defect (inherited coagulopathy) in the WAG/RijYcb rats.

The factors responsible for clotting were evaluated and properly assessed for their activity. The animals harboring the mutation were affected and on evaluation of individual clotting factor activities it was found that the affected animals were deficient in a specific factor VIII (FVIII). The FVIII gene is located on the autosome (chromosome number 18) in rats whereas in human and mice it is located on the X chromosome. When the factor VIII cDNA was sequenced it was found that a point mutation, resulting in a substitution of Leucine at 176 th position by Proline (L176P), in the A1 domain of the factor VIII protein had occurred. This mutation is responsible for the disruption of the tertiary structure of the FVIII protein molecule.

When the affected animals were administered with the human plasma or human recombinant FVIII it rectified the coagulation defects.

From this study it can be concluded that this model is unique because the size of the rat is larger as compared to mice and this coagulation defect is present in both sexes. This model can be effectively used to study and develop new therapies for both hereditary as well as the acquired factor VIII deficiency.