Clinical, Biochemical, and Molecular Aspects of Glanzmann Thrombasthenia in Horses
Type of DegreeDissertation
MetadataShow full item record
Glanzmann thrombasthenia (GT) is an autosomal recessive bleeding disorder that is caused by a quantitative or qualitative deficiency in platelet membrane glycoprotein complex IIb-IIIa (integrin aIIbß3), also known as the fibrinogen receptor. Mutations in either of the genes encoding the aIIb or ß3 subunit can cause GT. This disease has been documented at the clinical and molecular levels in both human beings and dogs. In our study, two horses were examined that had a history of chronic epistaxis. Horse 1 was a seven-year old Thoroughbred-cross gelding located in England, while horse 2 was a four-year old American Quarter Horse mare located in Auburn, Alabama. Initial findings for both included normal coagulation screening test results, normal platelet numbers, and normal vWF antigen levels. Clot retraction and platelet aggregation responses to ADP, collagen, platelet activating factor and thrombin were impaired. Flow cytometric studies indicated a reduction in the aIIbß3 complex on platelet surfaces. Based on these findings, a diagnosis of Glanzmann thrombasthenia was made for both horses. The next part of this study entailed the determination of normal cDNA sequences encoding the aIIb and ß3 subunits in healthy horses and comparing them to human and canine sequences and cDNA from the horses with GT. The cDNA sequence from both GT horses showed a single guanine to cytosine substitution at position 122 in exon 2 of the gene encoding aIIb. This change results in the substitution of a proline for an arginine and likely causes instability in that region of the protein. Analysis of genomic DNA showed that horse 1 was homozygous and horse 2 heterozygous for this mutation. Based on these findings and the mode of inheritance of GT, it was theorized that horse 2 was a compound heterozygote, and that the mutation in the second allele was causing a lack of mRNA synthesis or nonsense-mediated decay. Sequence of non-coding areas of the aIIb gene in horse 2 was then compared with gene sequence in normal horses. Results showed that the aIIb gene from horse 2 had one allele with a 10-base-pair deletion that included the splice site between exon 11 and intron 11. This change would be predicted to result in lack of splicing between exons 11 and 12 and the appearance of a premature stop codon 50 base pairs downstream, which likely triggers nonsense-mediated decay of the RNA. The dam of horse 2 was heterozygous for this mutation; the sire was heterozygous for the exon 2 mutation. The sire and dam were clinically normal. This study represents the first characterization of cDNA sequences encoding platelet aIIb and ß3 in normal horses and the identification of two distinct mutations causing GT in horses.