| ObjectiveTo improve the medical knowledge on Glanzmann Thrombasthenia and contribute to the existing literature,we constructed a new mutant protein structure model to predict the possible impact of new mutant genes on protein function,and analyzed the correlation between clinical phenotypes and genetic characteristics of 5 Glanzmann Thrombasthenia cases.Methods1.Clinical and laboratory data of 5 patients diagnosed with Glanzmann Thrombasthenia at the First Affiliated Hospital of Guangxi Medical University between January 2015-December 2020 were analyzed.Modified Buchanan and Adix bleeding scoring standards were adopted,platelet aggregation test was used to observe the platelet response to various aggregation inducers, coagulation index and bleeding time were studied,and flow cytometry was used to detect the expression of platelet membrane glycoprotein GPⅡb/Ⅲa (CD41/CD61).2.High-throughput gene sequencing technology was used to screen for GT-related mutant genes,Sanger first-generation sequencing method was used for genetic sequencing analysis in order to identify the types of genetic mutations.3.PyMOL software was used for protein molecular modeling in order to evaluate and deepen the understanding of the influence of the new mutation on protein structure.Results(1)A total of 5 patients with GT,4 females and 1 male were examined.The age of diagnosis was 3 years-9 years old.Among the 5 GT patients,proband 1had a positive family history of bleeding while the rest of the patients had a negative family history of bleeding.Proband 2’s younger sister(patient 1)was diagnosed as a GT patient.All patients were born in Guangxi Zhuang Autonomous Region.(2)The 5 patients had a history of bleeding of varying degrees.Probands1-4 had a grade 3 bleeding score,classified as high-risk moderate bleeding for probands 1 and 2 and low-risk moderate bleeding for probands 3 and 4.Proband2’s younger sister(patient 1)had a level 2 bleeding score,classified as mild bleeding.(3)All coagulation parameters and platelet counts in the 5 patients were normal,but the bleeding time was prolonged with severely reduced platelets response to various aggregation inducers such as adenosine diphosphate,thrombin,epinephrine,collagen,arachidonic acid,etc.,while response to ristomycin was basically normal.The results of flow cytometry showed that probands 1,2,and patient1 had type III GT while probands 3 and 4 had type I GT.(4)Gene sequencing showed compound heterozygous mutations ITGB3 p.Cys549Ser/p.Leu705 Cysfs Ter4 in proband 1,compound heterozygous mutations ITGB3 p.Cys549Ser/p.Gln254 Lys in proband 2 and her sister,compound heterozygous mutations ITGA2 B p.Leu973 Alafs Ter63/p.Glu795 Lysfs Ter31 in proband 3 and homozygous mutation ITGB3 Gln327 Ter in proband 4.Heterozygosity at the corresponding mutation sites was detected in some members of the probands families.Mutations Gln254 Lys,Glu795Lysfs Ter31,and Gln327 Ter are reported for the first time.(5)Spatial structural analysis of the mutant protein showed that ITGB3Gln254 Lys mutation had changed the intermolecular force in the βA domain,and affected the normal binding of the GPIIb/IIIa complex,which may result in the functional damage of the GPIIb/IIIa molecules.Conclusion:1.The clinical manifestations of patients with Glanzmann Thrombasthenia are heterogeneous.There is no clear correlation between clinical bleeding severity and the expression level of GPⅡb/Ⅲa complex.It is possible that GPⅡb/Ⅲa functional defects are more prone to lead to bleeding events than insufficient synthesis.2.ITGB3 Cys549Ser/Leu705 Cysfs Ter4 compound heterozygous mutation and ITGB3 Cys549Ser/Gln254 Lys compound heterozygous mutation can lead to type III GT,ITGA2 B Leu973Alafs Ter63/Glu795 Lysfs Ter31 compound heterozygous mutation and ITGB3 Gln327 Ter homozygous mutation can cause type I GT.3.ITGB3 Gln254 Lys,ITGB3 Gln327 Ter and ITGA2 B Glu795Lysfs Ter31 mutations have not been reported before.These may be new mutant genotypes causing GT. |
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