SABIIA - Sistema Aberto e Integrado de Informação em Agricultura (Sabiia)

Wenning,M.R.S.C.; Kimura,E.M.; Costa,F.F.; Saad,S.T.O.; Gervásio,S.; de Jorge,S.B.; Borges,E.; Silva,N.M.; Sonati,M.F..

Seven unrelated patients with hemoglobin (Hb) H disease and 27 individuals with alpha-chain structural alterations were studied to identify the alpha-globin gene mutations present in the population of Southeast Brazil. The -alpha3.7, --MED and -(alpha)20.5 deletions were investigated by PCR, whereas non-deletional alpha-thalassemia (alphaHphalpha, alphaNcoIalpha, aaNcoI, alphaIcalpha and alphaTSaudialpha) was screened with restriction enzymes and by nested PCR. Structural alterations were identified by direct DNA sequencing. Of the seven patients with Hb H disease, all of Italian descent, two had the -(alpha)20.5/-alpha3.7 genotype, one had the --MED/-alpha3.7 genotype, one had the --MED/alphaHphalpha genotype and...

Characterization of alpha thalassemic genotypes by multiplex ligation-dependent probe amplification in the Brazilian population

Palavras-chave: Alpha-globin genes; Alpha-globin structural variants; Alpha-thalassemia; Hemoglobin H; Hb H disease; Hemoglobin variants; Hemoglobinopathies.

Ano: 2000

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2000000900008

Suemasu,C.N.; Kimura,E.M.; Oliveira,D.M.; Bezerra,M.A.C.; Araújo,A.S.; Costa,F.F.; Sonati,M.F..

Alpha-thalassemia is the most common inherited disorder of hemoglobin synthesis. Genomic deletions involving the alpha-globin gene cluster on chromosome 16p13.3 are the most frequent molecular causes of the disease. Although common deletions can be detected by a single multiplex gap-PCR, the rare and novel deletions depend on more laborious techniques for their identification. The multiplex ligation-dependent probe amplification (MLPA) technique has recently been used for this purpose and was successfully used in the present study to detect the molecular alterations responsible for the alpha-thalassemic phenotypes in 8 unrelated individuals (3 males and 5 females; age, 4 months to 30 years) in whom the molecular basis of the disease could not be determined...

Death switch for gene therapy: application to erythropoietin transgene expression

Palavras-chave: Alpha-thalassemia; Hb H disease; Multiplex ligation-dependent probe amplification; Genetic polymorphisms; Brazilian population.

Ano: 2011

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2011000100003

Souza,D.S.; Spencer,D.M.; Salles,T.S.I.; Salomão,M.A.; Payen,E.; Beuzard,Y.; Carvalho,H.F.; Costa,F.F.; Saad,S.T.Olalla.

The effectiveness of the caspase-9-based artificial "death switch" as a safety measure for gene therapy based on the erythropoietin (Epo) hormone was tested in vitro and in vivo using the chemical inducer of dimerization, AP20187. Plasmids encoding the dimeric murine Epo, the tetracycline-controlled transactivator and inducible caspase 9 (ptet-mEpoD, ptet-tTAk and pSH1/Sn-E-Fv’-Fvls-casp9-E, respectively) were used in this study. AP20187 induced apoptosis of iCasp9-modified C2C12 myoblasts. In vivo, two groups of male C57BI/6 mice, 8-12 weeks old, were injected intramuscularly with 5 µg/50 g ptet-mEpoD and 0.5 µg/50 g ptet-tTAk. There were 20 animals in group 1 and 36 animals in group 2. Animals from group 2 were also injected with the 6 µg/50 g iCasp9...

Detection of cytomegalovirus infections by PCR in renal transplant patients

Palavras-chave: Gene therapy; Erythropoietin; Death switch; Caspase 9; AP20187; Anemia.

Ano: 2010

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000700005

Costa,S.C.B.; Miranda,S.R.P.; Alves,G.; Rossi,C.L.; Figueiredo,L.T.M.; Costa,F.F..

Cytomegalovirus (CMV) is the single most important infectious agent affecting recipients of organ transplants. To evaluate the incidence and the clinical importance of CMV infection in renal transplants in Brazil, 37 patients submitted to renal allograft transplants were tested periodically for the presence of cytomegalovirus DNA in urine using the polymerase chain reaction (PCR), and for the presence of IgM and IgG antibodies against CMV by enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescence (IIF). The PCR-amplified products were detected by gel electrophoresis and confirmed by dot-blot hybridization with oligonucleotide probes. Thirty-two of the 37 patients (86.4%) were positive by at least one of the three methods. In six patients,...

Detection of somatic mutations of the PIG-A gene in Brazilian patients with paroxysmal nocturnal hemoglobinuria

Palavras-chave: Transplantation; CMV; PCR; Kidney.

Ano: 1999

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X1999000800004

Franco de Carvalho,R.; Arruda,V.R.; Saad,S.T.O.; Costa,F.F..

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal syndrome characterized by intravascular hemolysis mediated by complement, thrombotic events and alterations in hematopoiesis. Basically, the molecular events which underlie the complexity of the syndrome consist of the absence of the glycosylphosphatidylinositol (GPI) anchor as a consequence of somatic mutations in the PIG-A gene, located on the X chromosome. The GPI group is responsible for the attachment of many proteins to the cytoplasmic membrane. Two of them, CD55 and CD59, have a major role in the inhibition of the action of complement on the cellular membrane of blood cells. The absence of GPI biosynthesis can lead to PNH. Since mutations in the PIG-A gene are always present in patients...

Efeito da temperatura das amostras de leite na concentração de cálcio solúvel e de beta-caseína: interferência no teste de estabilidade frente ao etanol

Palavras-chave: Paroxysmal nocturnal hemoglobinuria; Intravascular hemolysis; Complement PIG-A; Mutation.

Ano: 2001

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2001000600010

Arq. Bras. Med. Vet. Zootec.

Costa,F.F.; Brito,M.A.V.P.; Souza,G.N.; Pereira,D.B.C.; Pinto,I.S.B.; Martins,M.F..

O objetivo deste trabalho foi avaliar a variação do perfil proteico e do cálcio solúvel na coagulação do leite pelo etanol nas temperaturas de 4ºC, 10ºC, 15ºC e 20ºC. Amostras de leite de 61 animais foram avaliadas quanto à estabilidade ao etanol nas concentrações de 66 a 92% (v/v) nas temperaturas de 4ºC, 10ºC, 15ºC e 20ºC. Três amostras, após 24 horas de armazenamento a 4ºC, foram ultracentrifugadas em quadruplicata (40.000 x g) a 4ºC e a 20ºC, respectivamente, por 60 minutos. Em seguida, o sobrenadante foi retirado e submetido à análise do cálcio solúvel pela técnica via úmida (digestão nitroperclórica) e leitura em espectrofotômetro de absorção atômica. O perfil proteico foi analisado pela técnica de eletroforese capilar empregando kit específico para...

Hemochromatosis (HFE) gene mutations in Brazilian chronic hemodialysis patients

Palavras-chave: Temperatura do leite; Etanol; Cálcio; Β-caseína; Eletroforese.

Ano: 2014

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0102-09352014000200034

Perícole,F.V.; Alves,M.A.V.R.; Saad,S.T.O.; Costa,F.F..

Patients with chronic renal insufficiency (CRI) have reduced hemoglobin levels, mostly as a result of decreased kidney production of erythropoietin, but the relation between renal insufficiency and the magnitude of hemoglobin reduction has not been well defined. Hereditary hemochromatosis is an inherited disorder of iron metabolism. The importance of the association of hemochromatosis with treatment for anemia among patients with CRI has not been well described. We analyzed the frequency of the C282Y and H63D mutations in the HFE gene in 201 Brazilian individuals with CRI undergoing hemodialysis. The analysis of the effects of HFE mutations on iron metabolism and anemia with biochemical parameters was possible in 118 patients of this study (hemoglobin,...

High prevalence of alpha-thalassemia among individuals with microcytosis and hypochromia without anemia

Palavras-chave: HFE; Chronic renal insufficiency; Anemia; Iron metabolism.

Ano: 2005

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2005000900005

Borges,E.; Wenning,M.R.S.C.; Kimura,E.M.; Gervásio,S.A.; Costa,F.F.; Sonati,M.F..

In order to determine the contribution of alpha-thalassemia to microcytosis and hypochromia, 339 adult outpatients seen at Unicamp University Hospital (with the exception of the Clinical Hematology outpatient clinics), who showed normal hemoglobin (Hb) levels and reduced mean corpuscular volume and mean corpuscular hemoglobin, were analyzed. Ninety-eight were Blacks (28.9%) and 241 were Caucasians (71.1%). In all cases, Hb A2 and F levels were either normal or low. The most common deletional and nondeletional forms of alpha-thalassemia [-alpha3.7, -alpha4.2, --MED, -(alpha)20.5, alphaHphIalpha, alphaNcoIalpha, aaNcoI and alphaTSAUDI] were investigated by PCR and restriction enzyme analyses. A total of 169 individuals...

Influence of the polymorphisms of the α-major regulatory element HS-40 on in vitro gene expression

Palavras-chave: Alpha-thalassemia; Microcytosis; Hypochromia; Hemoglobinopathies; Brazilian population.

Ano: 2001

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2001000600009

Ribeiro,D.M.; Zaccariotto,T.R.; Santos,M.N.N.; Costa,F.F.; Sonati,M.F..

The α-MRE is the major regulatory element responsible for the expression of human α-like globin genes. It is genetically polymorphic, and six different haplotypes, named A to F, have been identified in some population groups from Europe, Africa and Asia and in native Indians from two Brazilian Indian tribes. Most of the mutations that constitute the α-MRE haplotypes are located in flanking sequences of binding sites for nuclear factors. To our knowledge, there are no experimental studies evaluating whether such variability may influence the α-MRE enhancer activity. We analyzed and compared the expression of luciferase of nine constructs containing different α-MRE elements as enhancers. Genomic DNA samples from controls with A (wild-type α-MRE) and B...

Low expression of APAF-1XL in acute myeloid leukemia may be associated with the failure of remission induction therapy

Palavras-chave: Alpha-major regulatory element; HS-40; Alpha-globin genes; Genetic polymorphisms; Gene expression.

Ano: 2009

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2009000900002

Benites,B.D.; Fattori,A.; Hackel,C.; Lorand-Metze,I.; De Souza,C.A.; Schulz,E.; Costa,F.F.; Saad,S.T.O..

Apoptotic protease activating factor 1 (APAF-1) has a critical role in the regulation of apoptosis. In the present study, the mRNA expression analysis of different APAF-1 transcripts (APAF-1S, APAF-1LC, APAF-1LN, and APAF-1XL) was analyzed in bone marrow samples from 37 patients with acute myeloid leukemia (newly diagnosed, with no previous treatment). APAF-1XL and APAF-1LN transcripts (with and without an extra WD-40 repeat region, respectively) were detected in all samples, although the major form expressed was APAF-1XL in 65% of the samples (group 1), while 35% of the samples expressed primarily APAF-1LN (group 2). Only 46% of the patients presented complete remission in response to remission induction therapy (represented by less than 5% marrow blasts...

Minimal doses of hydroxyurea for sickle cell disease

Palavras-chave: APAF-1; Acute myeloid leukemia; Apoptosis; Chemotherapy resistance.

Ano: 2008

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2008000700004

Lima,C.S.P.; Arruda,V.R.; Costa,F.F.; Saad,S.T.O..

The use of hydroxyurea (HU) can improve the clinical course of sickle cell disease. However, several features of HU treatment remain unclear, including the predictability of drug response and determination of adequate doses, considering positive responses and minimal side effects. In order to identify adequate doses of HU for treatment of sickle cell disease, 10 patients, 8 with sickle cell anemia and 2 with Sß thalassemia (8SS, 2Sß), were studied for a period of 6 to 19 months in an open label dose escalation trial (10 to 20 mg kg-1 day-1). Hemoglobin (Hb), fetal hemoglobin (Hb F) and mean corpuscular volume (MCV) values and reticulocyte, neutrophil and platelet counts were performed every two weeks during the increase of the HU dose and every 4 weeks...

Molecular identification of Sicilian (dß)º-thalassemia associated with ß-thalassemia and hemoglobin S in Brazil

Palavras-chave: Sickle cell disease; Hydroxyurea; Hemoglobinopathy.

Ano: 1997

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X1997000800004

Andrade,T.G. de; Fattori,A.; Saad,S.T.O.; Sonati,M.F.; Costa,F.F..

We describe the clinical and molecular characteristics of two unrelated Brazilian families with an association of the Sicilian form of (dß)º-thalassemia with hemoglobin S and ß-thalassemia. Direct sequencing of the ß-globin gene showed only the hemoglobin S mutation in patient 1 and the ß-thalassemia IVS1-110 in patient 2. The other allele was deleted in both patients and PCR of DNA samples of the breakpoint region of both patients showed a band of approximately 1,150 bp, expected to be observed in the DNA of carriers of Sicilian (dß)º-thalassemia. The nucleotide sequence of this fragment confirmed the Sicilian deletion. There are few reports concerning the Hb S/(dß)º-thalassemia association and patient 2 is the first reported case of Sicilian type of...

Presence of the RHD pseudogene and the hybrid RHD-CE-Ds gene in Brazilians with the D-negative phenotype

Palavras-chave: Thalassemia; Sickle cell; Hereditary persistence of fetal hemoglobin; Globin genes; Sicilian thalassemia.

Ano: 2002

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2002000800003

Rodrigues,A.; Rios,M.; Pellegrino Jr.,J.; Costa,F.F.; Castilho,L..

The molecular basis for RHD pseudogene or RHDpsi is a 37-bp insertion in exon 4 of RHD. This insertion, found in two-thirds of D-negative Africans, appears to introduce a stop codon at position 210. The hybrid RHD-CE-Ds, where the 3' end of exon 3 and exons 4 to 8 are derived from RHCE, is associated with the VS+V- phenotype, and leads to a D-negative phenotype in people of African origin. We determined whether Brazilian blood donors of heterogeneous ethnic origin had RHDpsi and RHD-CE-Ds. DNA from 206 blood donors were tested for RHDpsi by a multiplex PCR that detects RHD, RHDpsi and the C and c alleles of RHCE. The RHD genotype was determined by comparison of size of amplified products associated with the RHD gene in both intron 4 and exon 10/3'-UTR. VS...

Screening for mutations in human alpha-globin genes by nonradioactive single-strand conformation polymorphism

Palavras-chave: RHD pseudogene; RHD-CE-Ds; D-negative phenotype; VS antigen; Brazilians.

Ano: 2002

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2002000700002

Jorge,S.B.; Melo,M.B.; Costa,F.F.; Sonati,M.F..

Point mutations and small insertions or deletions in the human alpha-globin genes may produce alpha-chain structural variants and alpha-thalassemia. Mutations can be detected either by direct DNA sequencing or by screening methods, which select the mutated exon for sequencing. Although small (about 1 kb, 3 exons and 2 introns), the alpha-globin genes are duplicate (alpha2 and alpha1) and highy G-C rich, which makes them difficult to denature, reducing sequencing efficiency and causing frequent artifacts. We modified some conditions for PCR and electrophoresis in order to detect mutations in these genes employing nonradioactive single-strand conformation polymorphism (SSCP). Primers previously described by other authors for radioactive SSCP and phast-SSCP...

ß-Spectrin São PauloII, a novel frameshift mutation of the ß-spectrin gene associated with hereditary spherocytosis and instability of the mutant mRNA

Palavras-chave: Alpha-globin genes; Nonradioactive SSCP; Alpha-globin structural variants; Hemoglobinopathies; Mutation screening.

Ano: 2003

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2003001100004

Bassères,D.S.; Tavares,A.C.; Costa,F.F.; Saad,S.T.O..

Hereditary spherocytosis (HS) is a common inherited anemia characterized by the presence of spherocytic red cells. Defects in several membrane protein genes have been involved in the pathogenesis of HS. ß-Spectrin-related HS seems to be common. We report here a new mutation in the ß-spectrin gene coding region in a patient with hereditary spherocytosis. The patient presented acanthocytosis and spectrin deficiency and, at the DNA level, a novel frameshift mutation leading to HS, i.e., a C deletion at codon 1392 (ß-spectrin São PauloII), exon 20. The mRNA encoding ß-spectrin São PauloII was very unstable and the mutant protein was not detected in the membrane or in other cellular compartments. It is interesting to note that frameshift mutations of the...

Thalassemia intermedia as a result of heterozygosis for ß0-thalassemia and aaaanti-3.7/aa genotype in a Brazilian patient

Palavras-chave: SS-Spectrin; Red cell membrane; Spherocytosis; Acanthocytes; Mutation.

Ano: 2002

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2002000800009

Kimura,E.M.; Grignoli,C.R.E.; Pinheiro,V.R.P.; Costa,F.F.; Sonati,M.F..

We report a case in which the interaction of heterozygosis for both the ß0-IVS-II-1 (G->A) mutation and the aaaanti-3.7 allele was the probable cause for the clinical occurrence of thalassemia intermedia. The propositus, a 6-year-old Caucasian Brazilian boy of Portuguese descent, showed a moderately severe chronic anemia in spite of having the ß-thalassemia trait. Investigation of the alpha-globin gene status revealed heterozygosis for alpha-gene triplication (aaa/aa). The patient's father, also presenting mild microcytic and hypochromic anemia, had the same alpha and ß genotypes as his son, while the mother, not related to the father and...

The Agamma-195 (C->G) mutation in hereditary persistence of fetal hemoglobin is not associated with activation of a reporter gene in vitro

Palavras-chave: SS-Thalassemia; Thalassemia intermedia; Triplicated alpha-globin genes; Hemoglobinopathies.

Ano: 2003

URL: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2003000600003

Schreiber,R.; Gonçalves,M.S.; Junqueira,M.L.; Saad,S.T.O.; Krieger,J.E.; Costa,F.F..

Hereditary persistence of fetal hemoglobin is an uncommon, benign disorder in which the expression of gamma-globin genes persists into adult life. Several point mutations have been associated with the increased gamma-globin gene promoter activity. We evaluated the -195 (C->G) mutation by a functional in vitro assay based on the luciferase reporter gene system. The results indicated that the increased promoter activity observed in vivo could not be reproduced in vitro under the conditions employed, suggesting that other factors may be involved in the overexpression of the gamma-globin gene containing the -195 (C->G) mutation. Furthermore, this is the first time that the -195 (C->G) mutation of the Agamma-globin gene has been evaluated by in vitro...