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Biomedical subjects

A Iolascon

Publications and source records attributed to A Iolascon.

At least 109 records · Page 6Linked to original sources
PubMed1995

High frequency of homozygous deletions of CDK4I gene in childhood acute lymphoblastic leukaemia.

To determine the incidence of homozygous deletions of the newly identified tumour suppressor gene, CDK4I, molecular genomic DNA analyses by PCR technique were performed on primary neoplastic cells from 22 childhood acute leukaemias obtained at presentation. The blast cells derived in all the analysed cases from bone marrow. We found that none of acute myeloblastic leukaemias (four cases) showed the CDK4I alteration, whereas 6/13 (46%) common acute lymphoblastic leukaemias (ALLs) displayed homozygous deletions. Moreover, and even more important, all the blasts purified from ALLs derived from early lymphoid precursors (three early-T ALLs and two pre-B ALLs) showed the absence of CDK4I gene. When the entire coding sequence of the CDK4I gene from samples without homozygous deletions was analysed by the single-strand conformational polymorphism method, no point mutations were identified. These results demonstrate that CDK4I gene deletions are very frequent and probably early events in childhood acute leukaemias of lymphoid origin and especially in early-T and pre-B ALLs. Moreover, the molecular mechanism of the loss of function of the gene is correlated, at least in childhood ALLs, almost exclusively to deletions and not to point mutations.

Base Sequence↗
PubMed1995

Congenital dyserythropoietic anemia type II associated with G6PD Seattle in a Sicilian child.

A 2-year-old Sicilian boy was investigated because of chronic nonspherocytic hemolytic anemia (CNSHA) associated with hepatosplenomegaly. Appropriate studies revealed deficiency of glucose-6-phosphate dehydrogenase type Seattle (G6PD Seattle). In addition, bone marrow morphology, serological studies and analysis of red cell membrane proteins revealed congenital dyserythropoietic anemia (CDA) type II (or HEMPAS). Because G6PD Seattle on its own does not cause CNSHA, we believe that the clinical manifestations in this patient are essentially due to the CDA type II abnormality. However, the coexistence of these two different red cell abnormalities may affect the clinical picture specifically by making CDA type II more hemolytic than it would have been otherwise.

Anemia, Dyserythropoietic, Congenital↗
PubMed1995

Spectrin Anastasia (alpha I/78): a new spectrin variant (alpha 45 Arg-->Thr) with moderate elliptocytogenic potential.

We describe a white Italian kindred in which hereditary elliptocytosis (HE) is associated with abnormal level of alpha I/78 peptide in spectrin digest. Clinical phenotype varied among the family members ranging from asymptomatic to mild haemolytic HE. The original mutation responsible is a G-C substitution of the spectrin alpha-gene: alpha 45 Arg-->Thr (AGG-->ACG). The corresponding spectrin is designated spectrin Anastasia. Utilizing a secondary structure predictive method we suggest that this mutation has a poor capability to induce conformational changes of the tetramerization site and thus shows a moderate elliptocytogenic potential.

Adult↗
PubMed1994

Mild elliptocytosis associated with the alpha 34 Arg-->Trp mutation in spectrin Genova (alpha I/74).

We report a new mutation responsible for nonhemolytic hereditary elliptocytosis (HE). The proband displayed an impaired spectrin self-association and an increase of the alpha I 74-kD fragment (alpha I/74 abnormality). The responsible mutation occurred in exon 2 of spectrin alpha-gene: alpha 34 Arg-->Trp (CGG-->TGG), defining spectrin Genova. In Trans to allele alpha Genova, the proband disclosed allele alpha LELY, a common low-expression allele of spectrin alpha-gene. It was recognized through particular peptide maps as well as characteristic mutations in exon 40 and intron 45, respectively. The father, who carried allele alpha Genova, but not allele alpha LELY, had a milder presentation. The sensitization of allele alpha Genova by allele alpha LELY was noticeable in the proband as compared with his father. Nevertheless, it was not as sharp as that observed with many other alpha I/74 HE alleles. Therefore, each alpha I/74 HE allele has a distinct intrinsic severity.

Alleles↗
PubMed1994

Maple syrup urine disease (MSUD): screening for known mutations in Italian patients.

Maple syrup urine disease (MSUD) is an autosomal recessive disease due to deficiency of the branched-chain alpha-ketoacid dehydrogenase (BCKDH) caused by a large number of mutations. In the present study, DNA from Italian patients and their relatives was examined for three point mutations (Y393N in the E1 alpha gene, T841G and G1031A in the E2 gene) and two deletions (-G at the intron/exon border of exon 8 in the E2 gene and an 11 bp deletion in exon 1 of the E1 beta gene) using the polymerase chain reaction (PCR) followed by allele-specific oligonucleotide (ASO) hybridization, gene-scanning size analysis of fluorescent-tagged PCR products and/or automated DNA sequence analysis. Our results show that two different mutations account for 7 of the 20 mutant MSUD alleles. Two unrelated affected children, two of their parents and one sibling were carriers for the 11 bp deletion in the E1 beta gene, one patient and her mother were heterozygous for Y393N in E1 alpha, while T841G, G1031A and the -G deletion in E2 were not detected. This study is the first attempt to characterize at a nucleic acid level MSUD mutations in Italy. Our results indicate that additional defects are present in the Italian population and that, unlike the Mennonites, a number of different MSUD mutations exist in Italians.

Alleles↗
PubMed1994

Glucose 6-phosphate dehydrogenase deficiency and red cell membrane defects: additive or synergistic interaction in producing chronic haemolytic anaemia.

We have investigated two unrelated patients with congenital haemolytic anaemia in both of whom we found a combination of hereditary spherocytosis (HS) and glucose-6-phosphate dehydrogenase (G6PD) deficiency. Segregation of the two defects was documented in both families, who had different molecular abnormalities for both HS and G6PD deficiency. In one family the propositus had a reduced level of spectrin and G6PD Seattle (282Asp-->His). In the other family the propositus had a band 3 abnormality and was heterozygous for G6PD Mediterranean (188Ser-->Phe). From a comparison of clinical and haematological findings in family members with either or both abnormalities we conclude that in one case the two defects exhibited a synergistic effect, resulting in a severe chronic haemolytic anaemia; whereas in the other the association was simply additive.

Anemia, Hemolytic, Congenital↗
PubMed1994

Erythrocyte membrane protein alterations underlying clinical heterogeneity in hereditary spherocytosis.

Hereditary spherocytosis (HS) is a very heterogenous condition both at clinical and biochemical level. To establish the relationship between these aspects we performed a clinical and biochemical study in 87 Italian HS subjects. Patients were divided into three groups based on clinical severity (mild, typical and severe) and into five subgroups based on specific membrane abnormalities identified by polyacrylamide gel electrophoresis (isolated spectrin deficiency, spectrin deficiency combined with mild ankyrin reduction, spectrin deficiency combined with severe ankyrin reduction, band 3 reduction and isolated protein 4.2 reduction). We were not able to assess any alteration in six HS patients. A good correlation between clinical HS forms and membrane protein defects is shown. We conclude that erythrocyte membrane analysis should be carried out after diagnosis of HS in order to predict the clinical course of the disease.

Adolescent↗
PubMed1994

A deletional frameshift mutation in spectrin beta-gene associated with hereditary elliptocytosis in spectrin Napoli.

We studied a clinically manifest, dominantly transmitted elliptocytosis in an Italian family. We found a new spectrin variant, designated spectrin Napoli. Its beta-chain was truncated in its C-terminal region (apparent MW 216 kD). It displayed a low expression level (15%). There was a 8 nt deletion: CTTTTGAGAAGT-->CTGT (nt 6255-6262), starting after codon 2053. This deletion was followed by a 54 nt (18 amino acids) missense sequence and terminated by the TGA triplet which normally overlaps codons 2074 and 2075 (CTTGAG). The overall length of the mutated beta-chain was comparable to that found in spectrin Nice, spectrin Tokyo and spectrin Tandil, which are other variants with truncated beta-chains; however, a distinct nonsense codon was used in spectrin Napoli.

Adult↗
PubMed1994

Rapid detection of medium chain acyl-CoA dehydrogenase gene mutations by non-radioactive, single strand conformation polymorphism minigels.

Medium chain acyl-CoA dehydrogenase (MCAD) deficiency is a common inherited metabolic disorder affecting fatty acid beta oxidation. Identification of carriers is important since the disease can be fatal and is readily treatable once diagnosed. Twelve molecular defects have been identified in the MCAD gene; however, a single highly prevalent mutation, A985G, accounts for > 90% of mutant alleles in the white population. In order to facilitate the molecular diagnosis of MCAD deficiency, oligonucleotide primers were designed to amplify the exon regions encompassing the 12 mutations enzymatically, and PCR products were then screened with a single strand conformation polymorphism (SSCP) based method. Minigels were used allowing much faster run times, and silver staining was used after gel electrophoresis to eliminate the need for radioisotopic labelling strategies. Our non-radioactive, minigel SSCP approach showed that normals can be readily distinguished from heterozygotes and homozygotes for all three of the 12 known MCAD mutations which were detected in our sampling of 48 persons. In addition, each band pattern is characteristic for a specific mutation, including those mapping in the same PCR product like A985G and T1124C. When necessary, the molecular defect was confirmed using either restriction enzyme digestion of PCR products or by direct DNA sequence analysis or both. This rapid, non-radioactive approach can become routine for molecular diagnosis of MCAD deficiency and other genetic disorders.

Acyl-CoA Dehydrogenase↗
PubMed1993

Spectrin cagliari. an Ala-->Gly substitution in helix 1 of beta spectrin repeat 17 that severely disrupts the structure and self-association of the erythrocyte spectrin heterodimer.

The spectrin tetramer, the principal structural element of the red cell membrane skeleton, is formed by stable head-to-head self-association of two spectrin heterodimers. The self-association site appears to be formed by interactions between helices 1 and 2 of beta spectrin repeat 17 of one dimer with helix 3 of alpha spectrin repeat 1 of the other dimer to form two combined alpha-beta triple-helical segments. The head of the heterodimer appears to involve similar intradimer interactions. We describe the first example of an amino acid substitution in helix 1 of this combined alpha-beta triple-helical segment, which, although relatively minor, profoundly impairs tetramer formation. Strikingly, low angle rotary shadowing electron microscopy of isolated spectrin dimers reveals that this mutation also severely disrupts the head of the heterodimer causing it to be open. Following linkage studies which were most consistent with a beta spectrin gene mutation, a nucleotide change was identified in codon 2018, resulting in an Ala-->Gly substitution in the first helical domain of beta spectrin repeat 17. Because glycine is a strong helix breaker, this change is predicted to disrupt the conformation of this helical domain. Our results indicate that this helical domain must play direct roles in the alpha-beta interdimer interactions that form the self-association site of the tetramer and in the alpha-beta intradimer interactions at the head of the heterodimer.

Alanine↗
PubMed1993

Coexistence of hereditary spherocytosis (HS) due to band 3 deficiency and beta-thalassaemia trait: partial correction of HS phenotype.

A kindred with hereditary spherocytosis and beta-thalassaemia trait was identified. Detailed studies of the red cell membrane proteins on polyacrylamide gels with sodium dodecyl sulphate (SDS-PAGE) demonstrated the presence of band 3 (anion transporter) deficiency in all HS subjects (20-25% reduction) whereas spectrin content was in the normal range. The molecular defect of beta thalassaemia in this kindred was due to a beta(0) codon 39 (C-T) mutation, as assessed by beta globin gene amplification and ASO-probe hybridization. Seven subjects of this family were studied: two were normal, two had HS alone, two co-inherited HS and beta-thalassaemia trait, and one had beta-thalassaemia trait only. The two subjects with HS alone had a typical clinical form of spherocytosis with anaemia, reticulocytosis and increased red cell osmotic fragility. The two with both HS and beta-thalassaemia trait were not anaemic and showed a small, well-compensated haemolysis. Hence the finding of red cells with abnormalities of both HS and beta-thalassaemia indicates that beta-thalassaemic trait 'silences' HS caused by band 3 deficiency.

Adult↗
PubMed1993

Severe poikilocytosis associated with a de novo alpha 28 Arg-->Cys mutation in spectrin.

Severe poikilocytosis was observed in an Italian child. The mutation responsible was a de novo alpha 28 Arg-->Cys substitution (CGT-->TGT) in spectrin, a mutation known to cause hereditary elliptocytosis or hereditary pyropoikilocytosis. In this particular case the severity of the manifestations were accounted for by the occurrence, in trans to the alpha 28 mutation, of the alpha V/41 polymorphism. The latter has been shown previously to be associated with structural abnormalities at the alpha IV-alpha V domain junction and with a low expression level. The pronounced alteration of the dimer self association process was also explained by the location of the alpha 28 mutation. This mutation occurs in helix 3 of repeating segment alpha 1, e.g. precisely in the head-to-head contact between the spectrin alpha and beta chains. The present phenotype was compared to that yielded by another alpha 28 mutation (Arg-->His) also combined, in trans, with the alpha V/41 polymorphism. The pictures were very much alike, stressing the functional importance of residue alpha 28. The de novo character of the present mutation strengthens the view that codon alpha 28 is a 'hot spot' for mutations.

Arginine↗
PubMed1993

Atypical hereditary ovalocytosis associated with defective dyserythropoietic anemia.

The cases of a child and his mother affected by chronic anemia with atypical elliptocytosis are reported. When adolescent the mother underwent splenectomy, with an incomplete response. Anemia was characterized by a morphological picture of ovalocytosis associated with a significant percentage of spherocytes in the peripheral blood of the child and spiculated red cells in that of the splenectomized mother. Bone marrow aspirates of the child showed a striking erythropoietic hyperplasia with marked decrease of mature cells and dyserythropoietic features. Reticulocyte count was rather low. Ferrokinetics showed ineffective erythropoiesis. Biochemical studies on red blood cell membrane cytoskeleton showed that beta-spectrin, alpha-spectrin and protein 4.1, which are usually altered in hereditary elliptocytosis (HE), were normal in our cases. This report confirms the hypothesis of Torlontano who postulated the existence of a distinct atypical form of HE associated with ineffective and dysplastic erythropoiesis.

Anemia, Dyserythropoietic, Congenital↗
PubMed1993

p53 gene mutations in medulloblastoma. Immunohistochemistry, gel shift analysis, and sequencing.

Medulloblastoma (MB), the most common malignant tumor of the CNS in children, bears a loss of the short arm of chromosome 17 in almost half of the cases. The tumor suppressor gene p53 is located on this chromosome and its role in the pathogenesis of this primitive tumor is controversial. Twenty-two MBs were analyzed by single-strand conformation polymorphism (SSCP) of polymerase chain reaction-amplified conserved exons. Fragments displaying a gel mobility shift were subsequently analyzed by direct sequencing. Immunohistochemistry for p53 was performed in all cases; three had cytogenetic analysis. Two cases (9%) were found to harbor a mutation: one homozygous and one heterozygous. The latter showed focal p53 immunostaining. None of the cases with chromosome 17p abnormality by cytogenetic analysis were found to have a mutation in the remaining allele. Loss of heterozygosity (LOH) of 17p, however, was found in four cases (one by SSCP and three by cytogenetic analysis). Together with the homozygous deletion in one case, the overall incidence of p53 allelic involvement in MB is 23%. Although LOH for the p53 gene may confer a selective advantage to tumor cells harboring mutations with dominant negative oncogenic effect, the infrequent occurrence of p53 mutations in face of frequent LOH for this gene supports the previously formulated hypothesis of a novel tumor-related locus distal to p53 on chromosome 17p.

Adolescent↗
PubMed1992

Alpha I/65 hereditary elliptocytosis in southern Italy: evidence for an African origin.

alpha I/65 Hereditary elliptocytosis (HE) is due to the duplication of TTG codon 154 (leucine) of alpha-spectrin and is associated with a constant haplotype. It was encountered exclusively in African and American Blacks, and in North Africans. We assumed that it diffused from the Benin-Togo area to Northern Africa. We now report two South Italian families with alpha I/65 HE. The phenotype fully conformed to previous descriptions. The mode of transmission was dominant; however, the manifestations were more pronounced when the common, low expression level alpha V/41 allele occurred in trans to the alpha I/65 allele, also conforming to previous records. The mutation underlying alpha I/65 HE turned out to be, again, the duplication of TTG codon 154 and the associated haplotype was the same as that encountered previously (+-+; XbaI, PvuII, MspI). Thus, the alpha I/65 allele found in Italy must have been introduced from North Africa across the Sicilian channel and would ultimately have originated from the Benin-Togo area. It would witness the same migratory stream as that followed by the Benin type haemoglobin S allele, which is also present in Southern Italy.

Africa, Western↗
PubMed1992

Spectrin/band 3 ratio as diagnostic tool in hereditary spherocytosis.

Recently it has been clearly established that partial deficiency of spectrin (SP) evaluated by radioimmunoassay is a common feature of many different forms of hereditary spherocytosis (HS). In this paper the determination of SP density (spectrin/band 3 ratio) in 46 HS patients and in 50 normal controls is presented. The comparison between the membrane SP density of HS subjects and controls showed a statistically significant difference (P less than 0.0005). Moreover no overlap between normal and HS subjects was observed. Membrane spectrin/band 3 ratio has been found related to some clinical features: indeed patients with severe HS showed a smaller SP density than those with milder HS. Our results show that the evaluation of membrane SP density permits a prompt diagnosis of HS and avoids extensive and unnecessary studies for other anaemias.

Adolescent↗