Study Of Clinical Phenotype And Mechanism In Twenty-five Chinese Pedigrees With Glanzmann Thrombasthenia

Glanzmann is an autosomal recessive bleeding syndrome affecting the megakaryocyte lineage and characterized by a quantitative or qualitative abnormality ofαIIbβ3, absent or severely reduced platelet aggregation in response to the physiological agonists including adenosine 5'-diphosphate (ADP), epinephrine, and collagen, and a relatively normal initial phase of aggregation in response to ristocetin in vitro (17). Most GT pathogenesis ascribes to the mutations of integrinαIIbβ3 gene, including deletions, insertions, inversions, single nucleotide substitutions that lead to missense or nonsense mutations, and splicing defects (18–22). IntegrinαIIbβ3, a calcium- dependent heterodimeric complex, is a prototype integrin that functions as a fundamental receptor for fibrinogen, VWF, fibronectin, vitronectin and CD40L, which plays a considerable role in normal thrombosis and hemostasis.Patients with Glanzmann's Thrombasthenia were often diagnosed about ten years old, usually by unexplained spontaneous mucosal bleeding. Skin petechia and petechiae was also very common, but not fatal. In our study, the age of onset of symptoms ranged from 0 to 7 years (mean 2 years), but the age at definitive diagnosis ranged from 1 to 29 years (median11 years). Twenty-five GT patients almost always had anemia ranging from mild to severe for chronic losing, but that was not the reason for patient's visits. They had bleeding tendency since childhood. Patients complaints concentrated mainly in mucocutaneous petechia and petechiae(25/25), epistaxis(9/25), gastrointestinal bleeding(3/25), gum bleeding(9/25), embryonic bleeding(6/25), menorrhagia(6/6), cerebral hemorrhage(1/25). No case appeared muscle haematoma and haemarthrosis.The maximum aggregation of platelet in response to the physiological agonists adenosine 5﹐-diphosphate (ADP) was lower significantly (range 0~18.2, mean 4.38)in vitro, but it's aggregation to ristocetin is normal(range 40.2~78.4, mean 55.9). Platelet surface expression ofαIIbβ3 was < 10%, 10%–50%, and > 50% of controls in thirteen, eight, and four patients, respectively. Platelet surface expression ofαIb was normal (range 61.5%~104%, mean 93.1%). However platelet membrane surface expression of integrinαIIb decreased obviously (range 1.2%~100%, mean 21.8%), and the expression of integrinβ3 was lower significantly (range 1.1~100, mean 15.1). Three patients showed normal surfaceαIIbβ3 expression. The binding of fibrinogen to activated platelets of two variant GT is significantly reduced, the rate of which was 39.5%,23.1% to normal respectively in three variant GT patients (low-to-normal, dysfunctional). The aggregation of platelet in response to the physiological agonists and platelet membrane surface expression of integrin was not related with the severity and frequencey of bleeding, at the same time which did not play a role in the performance prediction of bleeding. The lower binding ability of fibrinogen to activated platelets of GT did not indicate severity and frequencey of bleeding.Immunoblotting of the platelet lysates showed noαIIb in 14 patients, and noβ3 in 18 patients, but there plateletαIIbβ3 expression was normal as controls in three patients. Immunoblot analysis of the platelet lysates revealed normal GPIb in 23 patients. The total of platelet membrane surface expression of integrin is consistent with platelet membrane surface expression of integrin and inconsistent with the severity of hemorrhage.In our study 40 candidate pathogenic mutations were confirmed in 25 patients, including 23 missense mutations, 10 nonsense mutations, 3 splicing site mutations, and 1 frame mutation for one nucleotide insertion. 25 novel candidate pathogenic mutations were confirmed in seventeen patients. The missense mutations included 23Pro>His, 219Gly>Ser, 447Ala>Asn, 502Lys>Asn, 957Ser>Leu, 271Ala>Gly, 724Met>Ile, 576Cys>Tyr, 720Leu>Pro, 957Ser>Leu, 677Glu>lys, 646Thr>Ile inαIIb and 716Gly>Val, 385Val>Glu, 106Val>Ile, 525Gly>Asp, 34Arg>Gln, 647Asp>Glu inβ3. Meanwhile, four nonsense mutations and four frameshift mutations were found for the first time. Three novel nonsense mutations were identified inαIIb gene, namely 891Gln>stop, 351Arg>stop, 23Pro>stop. There were four novel point mutation at the splicing site ofαIIb intron, namelyαIIb IVS7(+2)T>C,αIIb IVS9(-1)G>A,αIIbIVS23(-1)G>A. More comprehensive coverage can be found in reviews discussing the structure and function of the GPIIb/IIIa receptor complex andthe clinical and molecular basis of Glanzmann's thrombasthenia.