Clinical And Molecular Genetic Analysis Of Two Pedigree With Idiopathic Epileptic Syndromes

The idiopathic epilepsies are considered to be primarily genetic in origin. Recent advances have demonstrated that many idiopathic epilepsies are ion channelopathies. Most of these diseases are caused by abnormal function of ligand-and voltage gated ion channels caused by a genetic defect. Generalized epilepsy with febrile seizures plus (GEFS+) is an autosomal dominant familial syndrome with a complex seizure phenotype. It is caused by mutations in one of 3 voltage-gated sodium channel subunit genes (SCN1B, SCN1A, and SCN2A) and the GABA(A) receptor gamma2 subunit gene (GBRG2). Benign familial neonatal convulsion (BFNC) is a rare autosomal dominant disorder caused by mutations in KCNQ2 and KCNQ3, two genes encoding for potassium channel subunits.Objective We collected two large multigenerational families to study the clinical and genetic characteristics of a Chinese family with GEFS+ and BFNC. The clinical data of two families were analyzed. The blood samples were taken for genetic analysis. DNA was extracted from peripheral blood leukocytes using phenolchloroform method.Part I: Clinical and Molecular Genetic Analysis of GEFS+Methods We performed linkage analysis on this pedigree. Eleven microsatellite markers spanning the critical regions on chromosomes 19ql3.1, 2q24,and 5q31.1-q33.1 were genotyped. These markers included D19S414,D19S220,D19S902 for the 19q region, D2S156,D2S142,D2S2330,D2S335,D2S364 for the 2q region, D5S436,D5S422 and D5S400 for the 5q region. DNA from each samplewas amplified for the 11 markers. After polymerase chain reactions (PCR), then the length of the PCR products was judged in the Strategene Eagle Eye II automated gel image analyzer. The date of PCR products were analyzed using the software Genescan v311, Genetyper v3.7. After Mendelian checking, the eligible genotyping data were used for linkage analysis. LOD scores were calculated by using MLJNK program of LINKAGE v5.1. The LOD scores were calculated at combination rate 0.0, 0.1, 0.2, 0.3, 0.4. Mutation analysis of GABRG2 gene was made by means of polymerse chain reaction (PCR)-direct sequencing in the proband and his father.Results1. The clinical characteristics of GEFS+: Six cases in a total of 20 family members over four generations. The phenotypes comprised febrile seizures (FS) in 1, febrile seizures plus (FS+) in 3; FS+ with typical absence in 1, and one remaining individuals, no information was available. All cases had favorable outcome and normal develoment.2. The genetic linkage study: Among the 11 selected microsatellite markers, (1) One marker date (D2S335) was omitted due to failed genotyping. (2) Two-point LODscores were below zero for the makers D19S414,D2S142,D2S156,D2S2330,D5S422, and the maximum LOD scores at 0.0 were less than -2 for the makers D19S220,D19S902,D2S364. Thus the Linkage result showed no evidence that thediseases locus was linked to above selected markers. (3) The other two markers D5S436 and D5S400 that two-point LOD scores at 0.0 were between 0.5～0.8 (>-2, and <3), the Linkage result for the two markers showed to be proved in next step.3. Mutational analysis of GABRG2: A synonymous mutation (C.1420OT) was detected on exon 11. Our results indicate that genomic variations of GABRG2 are not likely to be substantially involved in the etiology of GEFS+ in this family.Conclusion Our study failed to provide evidence supporting a causal relation between the SCN1A, SCN1B, GABRG2 mutation in this family. These results indicated that GEFS+ showed genetic heterogeneity. Part II: Clinical and Molecular Genetic Analysis of BFNCMethods All familial members were studied by clinical examinations and Mutation analysis of KCNQ2 and KCNQ3 genes were made by means of polymerse chain reaction (PCR)-direct sequencing in the proband and his father.Results1. The clinical characteristics of BFNC: In this study, we presented a pedigree families and 10 (5 girls and 5 boys) with BFNC. Nine started the similar seizures in the second or third days of life and remited by approximately 4 months of age, only one had the first seizure in 2ys. The phenotype included generalized seizures and/or partial seizures in this family. Interictal EEG and neuroradiologic studies were normal in proband. No subjects had other forms of epilepsy later in the life, Psychomotor development were normal.2. Mutational screeing of KCNQ2 gene (in the proband): (1) A synonymous mutation (c.911C>T) on exon 6; (2) A missense mutation (c.2339A>C) on exon 17(SNP 17.9%); (3) A missense mutation (c.2867C>T) on exon 17; (4) In addition, exon-intron boundaries mutation of KCNQ2 gene was detected near exon 13, when compared to wild-type. (5) It is ongoing to check mutation on exon 8.3. The mutation analysis of KCNQ3 gene: A missense mutation (c.1454C>A) were found on exon 9 in the proband.Conclusion We reported here the above mutation results. In the future, we want to identify whether or not the results are responsible for the BFNC phenotype or SNP in the KCNQ2 and KCNQ3 gene.