Purpose
A 50-year-old female with excessive menstruation and severe epistaxis since childhood was studied by using phenotype and genetic diagnosis. She was diagnosed as Glanzmann thrombasthenia (GT), meanwhile, her family members with severe epistaxis were also studied. Mutation sites were detected in them. Bioinformatics software was used to explore the molecular pathogenesis which provide evidence for in vitro and in vivo experiments.
Methods
Platelet count and morphology, coagulation function and platelet aggregation were detected for phenotype diagnosis. Platelet membrane glycoproteins (GP) αIIb and β3 were measured by flow cytometry (FCM). Exons and their end 15bp introns of 89 genes associated with hemorrhage and thrombosis from proband were amplified by PCR and analyzed by direct DNA sequencing. Mutation site was measured by sanger sequencing in other family members. ClustalX-2.1-win software was used to analyze homology of nucleotide. PROVEAN and SIFT was used to analyze the risk of mutation site.
Results
The proband, along with her brother and nephew, had normal platelet account and morphology. Coagulation function showed normal while platelet aggregation in response to ADP and AA were reduced. However, they had normal platelet aggregation in response to ristocetin. FCM demonstrated that expression level of CD41 and CD61 were significantly reduced. ITGA2B c.1067T>C heterozygous missense mutation, which led to a Val356→Ala substitution in αIIb protein, existed in the three people. This mutation site was highly conserved among human, mouse, rat and zebrafish. PROVEAN prediction scored -3.48 and SIFT prediction scored 0.001. These results demonstrated ITGA2B p.Val356Ala was pathogenic.
Conclusion
Glanzmann thrombasthenia is a rare autosomal recessive bleeding disorder results from quantitative or qualitative defect of αIIbβ3 integrin which encoded by ITGA2B and ITGB3. The nature of GT mutations is highly variable and most cases are sporadic. In our study, ITGA2B c.1067T>C (Val356Ala) heterozygous missense mutation caused Glanzmann thrombasthenia and aroused obvious symptoms. And in general, homozygous mutation in ITGA2B were recognized to led to Glanzmann thrombasthenia while ITGA2B heterozygous mutation carriers showed mild symptoms. However, there are also some patients with gain-of-function in ITGA2B showed abnormal platelet function or amount. More experiments should be conducted to identify the function of this heterozygous mutation.
No relevant conflicts of interest to declare.
Author notes
Asterisk with author names denotes non-ASH members.
