title: [Molecular study of red cell pyruvate kinase deficiency in 15 patients with chronic hemolytic anemia. Group of Erythropathology of Hematology and hemotherapy Association of Spain (HHAS)].
abstract: BACKGROUND
  Identification of RBC pyruvate-kinase (PK) gene mutations by polymerase chain reaction (PCR) and single strand conformation polymorphism (SSCP) followed by PK gene sequencing in positive cases has been assessed and the results obtained with a preliminary study of 15 unrelated patients of Spanish origin are presented.
  
  
  METHODS
  Patients have been classified into two different groups: group 1, propositus (15 cases), and group 2, relatives of the patients included in group 1 (10 males and 5 females). In group 1, a PCR was followed by SSCP and sequencing, and in group 2, the PCR was followed by digestion with specific restriction endonucleases (PCR-ER).
  
  
  RESULTS
  Group 1: from 15 patients included in the study 2 were identified as homozygous, 4 as heterozygous and 9 as compound heterozygous. In this group, were identify 26 affected alleles with 11 different mutations: T1456 10 alleles (38.6%), T721 3 alleles (11.6%), A1010, C514, C1015 and T1223 2 alleles (7.7%), and C1070, A1291, T1508, A1595 y T1675 one allele. Relatives from 8 out of 15 patients from group 1 showed the following pattern: homozygous (one case), heterozygous (10 cases), compound heterozygous (2 cases) and normal (2 cases).
  
  
  CONCLUSIONS
  SSCP procedure followed by direct gene sequencing in positive cases is fast and simple enough to allow the identification of PK deficient variants, avoiding the need of biochemical characterisation of semipurified deficient enzyme, which is more cumbersome and time consuming. In addition, the PCR-ER method is a very useful tool for screening of the most frequent molecular variants, as well as, for the detection of the carrier condition of this enzymopathy (family studies).
authors.affiliation: Departamento de Hematología, Hospital Clínic i Provincial, Barcelona.
authors.forename: R
authors.initials: R
authors.lastname: Zarza
authors.forename: A
authors.initials: A
authors.lastname: Pujades
authors.forename: J
authors.initials: J
authors.lastname: García
authors.forename: R
authors.initials: R
authors.lastname: Alvarez
authors.forename: A
authors.initials: A
authors.lastname: Carrera
authors.forename: J
authors.initials: J
authors.lastname: Estella
authors.forename: M
authors.initials: M
authors.lastname: Morey
authors.forename: A
authors.initials: A
authors.lastname: Remacha
authors.forename: G
authors.initials: G
authors.lastname: Pérez-Lungmus
authors.forename: B
authors.initials: B
authors.lastname: Nomdedeu
authors.forename: J
authors.initials: J
authors.lastname: Sánchez
authors.forename: J
authors.initials: J
authors.lastname: Gastearena
authors.forename: E
authors.initials: E
authors.lastname: Bureo
authors.forename: J L
authors.initials: JL
authors.lastname: Vives Corrons
mesh_terms: D000596:Amino Acids / Q000235:genetics; D000743:Anemia, Hemolytic / Q000201:enzymology / Q000235:genetics*; D002908:Chronic Disease; D004920:Erythropoiesis / Q000235:genetics*; D005260:Female; D005838:Genotype; D006405:Hematology; D006801:Humans; D008297:Male; D009711:Nucleotides / Q000235:genetics; D017354:Point Mutation / Q000235:genetics; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D011770:Pyruvate Kinase / Q000172:deficiency* / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D012955:Societies, Medical; D013030:Spain
chemical_list: D000596:Amino Acids; D009711:Nucleotides; D011770:Pyruvate Kinase
vernacular_title: Estudio molecular del déficit de piruvatocinasa eritrocitaria en 15 pacientes con anemia hemolítica crónica. Grupo de Eritropatología de la Asociación Española de Hematología y Hemoterapia (AEHH).