Study on molecular pathogenesis for a case of Glanzmann′s thrombasthenia
10.13602/j.cnki.jcls.2019.09.09
- VernacularTitle:1例血小板无力症的分子发病机制研究
- Author:
Bingjing GAO
1
;
Yan GONG
1
;
Chenxue QU
1
;
Ran YOU
1
;
Linzi MIAO
1
;
Yao LU
1
;
Tao LI
1
Author Information
1. Department of Clinical Laboratory, Peking University First Hospital
- Publication Type:Journal Article
- Keywords:
ITGA2B;
ITGB3;
Glanzmann′s thrombasthenia;
gene mutation
- From:
Chinese Journal of Clinical Laboratory Science
2019;37(9):680-685
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the molecular pathogenesis for a patient with Glanzmann thrombasthenia (GT).
Methods:The peripheral blood of a patient with Glanzmann′s thrombasthenia was collected, and the genetic mutations were detected by gene sequencing technology. The mutant plasmids were prepared by PCR site-directed mutagenesis and transfected into CHO-K1 cells of Chinese hamster ovary to construct in vitro eukaryotic expression system. The expressions of αⅡb and β3 protein subunits in CHO-K1 cells were detected by western blot. The expression levels of αⅡb and β3 in cellular membrane and cytoplasm of CHO-K1 cells were detected by flow cytometry. The expression and distribution of αⅡb and β3 in CHO-K1 cells were observed by immunofluorescent labeling under microscope.
Results:This patient was diagnosed with type Ⅱ GT. Gene sequencing revealed two mutations in ITGB3 gene which has not been reported in the literature. ITGB3 c.1495 T>C missense mutation resulted in replacement of cysteine no.499 by arginine (p.C499R). ITGB3 c.1728 delC code shift mutation resulted in a change in the amino acid synthesis initiated by the β3 protein subunit serine no.577 and terminated by the 92nd amino acid following these changes. The results of western blotting showed that the synthesis and expression of primary structures of αⅡb and β3 were detectable in the lysates of mutant CHO-K1 cells. The results of flow cytometry showed that no expression of β3 on the surface and intracellular of mutant CHO-K1 cells was observed. Under fluorescence microscopy no distribution of β3 protein subunit was displayed in mutant CHO-K1 cells.
Conclusion:The mutation of ITGB3 c.1728 del C or ITGB3 c.1495 T>C should be relevant to the cause of GT in this patient. The mutation of ITGB3 c.1728 del C and ITGB3 c.1495 T>C seems not to affect the formation of the primary structure of β3 protein subunit, but did affect the formation of its high-level structure.