Marina Pavić

Inherited Prothrombotic Risk Factors in Children With Stroke, Transient Ischemic Attack, or Migraine

OBJECTIVE. The aim of this study was to investigate the prevalence and possible association of inherited prothrombotic risk factors in children with stroke, transient ischemic attack, or migraine. METHODS. We performed genotypic analysis for factor V G1691A, factor II G20210A, methylenetetrahydrofolate reductase C677T, and 4 common platelet glycoprotein polymorphisms (human platelet alloantigen-1, -2, -3, and -5) in 150 children <18 years of age with established diagnoses of stroke, transient ischemic attack, or migraine. Children were classified into 5 groups, namely, childhood arterial ischemic stroke (N = 33), perinatal arterial ischemic stroke (N = 26), hemorrhagic stroke (N = 20), transient ischemic attack (N = 36), and migraine (N = 35). The control group consisted of 112 children ≤18 years of age from the same geographical region who had no history of neurologic or thromboembolic diseases. RESULTS. Heterozygosity for factor V G1691A was associated with approximately sevenfold increased risk for arterial ischemic stroke, perinatal arterial ischemic stroke, and transient ischemic attack. Increased risk for transient ischemic attack was found in carriers of the human platelet alloantigen-2b allele, human platelet alloantigen-5a/b genotype, and combined human platelet alloantigen-2b and human platelet alloantigen-5b genotype. The presence of the human platelet alloantigen-2b allele was associated with a 2.23-fold increased risk for migraine, whereas carriers of the human platelet alloantigen-3b allele had a lower risk for arterial ischemic stroke than did carriers of the human platelet alloantigen-3a allele. CONCLUSIONS. Factor V G1691A has an important role in susceptibility to arterial ischemic stroke, both in the perinatal/neonatal period and in childhood, as well as transient ischemic attacks. A minor impact of human platelet alloantigen polymorphisms suggests that platelet glycoprotein polymorphisms may increase the risk of transient ischemic attacks and migraine, but this should be confirmed in larger studies.

Gene frequencies of platelet-specific antigens in Croatian population.

The human platelet antigens (HPA) are genetically defined polymorphisms expressed on platelet membrane glycoproteins. As platelet antigens are very important in several clinical situations and in population genetics, we used the polymerase chain reaction with sequence‐specific primers (PCR‐SSP) to investigate HPA‐1, ‐2, ‐3 and ‐5 allele frequencies in the Croatian population. The HPA frequencies obtained in 219 Croatians were: 1a–0·854, 1b–0·146, 2a–0·890, 2b–0·110, 3a–0·575, 3b–0·425, 5a–0·895 and 5b–0·105. These data are similar to the frequencies reported in most European studies with some significant differences in HPA‐2 when compared with the Dutch and German population, in HPA‐3 when compared with the Swiss population and in HPA‐5 when compared with the Finnish population. The three most common condensed HPA genotypes in the Croatian population were: HPA‐1a/a, ‐2a/a, ‐3a/b, ‐5‐a/a (0·283), HPA‐1a/a, ‐2a/a, ‐3a/a, ‐5‐a/a (0·137) and HPA‐1a/b, ‐2a/a, ‐3a/b, ‐5‐a/a (0·087). Data obtained in this study can be used for better understanding and treatment of immune‐mediated platelet disorders in our population.

Development and validation of a rapid method for genotyping three P-selectin gene polymorphisms based on high resolution melting analysis

High resolution melting (HRM) analysis is one of the newer, reliable, and sensitive genotyping techniques, which offers considerable time and cost savings. P‐selectin is an adhesion molecule that has a role in the initial phases of leukocyte adhesion to stimulated platelets and endothelial cells in inflammation. Multiple polymorphisms in P‐selectin gene (SELP) that affect the protein sequence have been described. The aim of this study was to design, optimize, and validate a simple and rapid in‐house HRM‐based method for genotyping the NM_003005.3:c.992G>A (c.992G>A), NM_003005.3:c.1918G>T (c.1918G>T), and NM_003005.3:c.2266A>C (c.2266A>C) SELP polymorphisms.

19 Inherited prothrombotic risk factors in children with perinatal arterial stroke

Perinatal arterial stroke (PAS), defined as a cerebrovascular event which occurs between 28 weeks of gestation and 28 days of postnatal age with either pathological or radiological evidence of focal arterial infarction, has received increased attention as an important cause of cerebral palsy and other neurologic disabilities, including epilepsy and cognitive impairment. Although sensitive neuroimaging techniques have dramatically improved the detection of PAS in recent years, the cause of PAS is still poorly understood. The increasing evidence that inherited or acquired prothrombotic disorders may be implicated in the pathogenesis of PAS prompted us to investigate the prevalence of genetic polymorphisms that encode proteins associated with thrombosis (factor V G1619A [FVL], factor II G20210A [PT], homocysteine metabolism (methylenetetrahydrofolate reductase C677T [MTHFR]), and human platelet antigens (HPA) in children with PAS. Polymorphisms were investigated in 24 children (11 boys, 13 girls) with PAS confirmed by brain imaging and in 103 children (72 boys, 31 girls) from the same geographical region that represented the control group, with standard laboratory techniques. At least 1 prothrombotic abnormality was identified in 18/24 (75 %) children with PAS: 3 were heterozygous for FVL, 16 were carriers of the MTHFR mutation (13 heterozygotes, 3 homozygotes) and 1 was double heterozygous for FVL and MTHFR ; the presence of PT 20210A was not detected. Similar genotype and allele frequencies of HPA-1, HPA-2 and HPA-5 were observed in both study groups. Although higher frequencies of HPA-3a/a (50%), HPA-3a allele (68.7%) and FVL (12.5%) were observed in children with PAS compared to the control group (28.2%, 53.4% and 1.9% respectively), these differences failed to reach statistical significance. On the contrary, homozygosity for MTHFR (12.5% in patients and 1.9% in controls) was found to be the only statistically significant difference between the studied groups.

Inherited prothrombotic risk factors in children with cerebrovascular disorders

Cerebrovascular disorders (CD) are an important cause of mortality and morbidity in children, and an emerging area for clinical research. There is increasing evidence that inherited or acquired prothrombotic disorders may be implicated in the etiology of CD in childhood. We have investigated the prevalence of genetic polymorphisms associated with thrombosis: factor V G1619A (FVL), factor II G20210A (PT), methylenetetrahidropholate reductase C677T (MTHFR), and human platelet antigens (HPA-1, -2, -3 and -5) in 151 children with CD and compared the data with those obtained for the control group (112 children from the same geographical region). Children with CD were classified into 5 groups according to diagnosis based on clinical symptoms, neurological examination, computed tomography and magnetic resonance imaging of the brain: children with arterial ischemic stroke (AIS ; n=36), perinatal arterial ischemic stroke (PAIS ; n=26), transient ischemic attack (TIA ; n=36), migraine (n=35) and intracranial hemorrhage (ICH ; n=18). Similar genotype distributions of all studied polymorphisms were found only between children with ICH and controls. Statistically significant differences between controls and other groups of children with CD were observed for at least one polymorphism: for FVL in children with AIS and TIA (p=0.048), for MTHFR in children with AIS (p=0.036), PAIS (p=0.034) and migraine (p=0.037), for HPA-2 and -5 (p=0.037) in children with TIA. Furthermore, higher frequencies of the HPA-3a allele in children with AIS were found to be statistically significant (p=0.027), as compared to the control group. Obtained results indicate that different polymorphisms are implicated in the etiology of CD in childhood.