Knowledge extraction using visualization of hemoglobin parameters to identify thalassemia

The analysis of large amounts of data is better performed by humans when represented in a graphical format. Therefore, a new research area called the Visual Data Mining is being developed endeavoring to use the number crunching power of computers to prepare data for visualization, allied to the ability of humans to interpret data presented graphically.This work presents the results of applying a visual data mining tool, called FastMapDB to detect the behavioral pattern exhibited by a dataset of clinical information about hemoglobinopathies known as thalassemia. FastMapDB is a visual data mining tool that get tabular data stored in a relational database such as dates, numbers and texts, and by considering them as points in a multidimensional space, maps them to a three-dimensional space. The intuitive three-dimensional representation of objects enables a data analyst to see the behavior of the characteristics from abnormal forms of hemoglobin, highlighting the differences when compared to data from a group without alteration.

Relationship between oxidative stress, glutathione S-transferase polymorphisms and hydroxyurea treatment in sickle cell anemia

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Serum melatonin level and oxidative stress in sickle cell anemia

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Rastreamento de hemoglobinas variantes e talassemias com associação de métodos de diagnóstico

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Diagnóstico laboratorial de hemoglobinas semelhantes à HbS

A hemoglobina S (HbS) está presente na população brasileira com prevalência variável, dependente dos grupos raciais formadores de cada região. A migração eletroforética em pH alcalino apresenta similaridade com outras hemoglobinas, e estudos complementares para sua correta caracterização são necessários. No presente estudo objetivamos traçar um fluxograma com as metodologias disponíveis para a caracterização da hemoglobina S e das hemoglobinas que apresentam migração semelhante em pH alcalino. No período de janeiro a junho de 2000, analisamos amostras de sangue com suspeita de hemoglobina S encaminhadas ao Laboratório de Hemoglobinas da Unesp. Caracterizamos diferentes mutantes e formas interativas com hemoglobina S, por procedimentos eletroforéticos, em variados pH, análises citológicas e testes bioquímicos específicos. Os procedimentos de análise aplicados resultaram em orientação fornecida aos laboratórios de rotina sobre como proceder no diagnóstico laboratorial destas alterações de hemoglobina. Desta forma contribuímos para um melhor conhecimento sobre a variabilidade genética das hemoglobinas em nossa população, auxiliando no acompanhamento clínico e no aconselhamento genético das hemoglobinopatias com fisiopatologia relacionada à alteração.

Prevalência de talassemias e hemoglobinas variantes no estado de Goiás, Brasil

As anemias hereditárias, em especial as talassemias e hemoglobinas (Hb) variantes, são as mais comuns das alterações genéticas humanas; sua freqüência na população brasileira é muito variável, dependendo dos grupos raciais formadores de cada região. O povoamento de Goiás, que teve início logo após o seu descobrimento, em 1726, motivado pela procura de ouro, foi composto principalmente por portugueses e escravos africanos, contexto que favoreceu a mestiçagem entre eles. Considerando que esses povos apresentam genes para as hemoglobinas anormais com freqüências variadas, é esperado que se encontrem essas alterações genéticas na nossa população. O objetivo deste trabalho foi avaliar a prevalência de talassemias e hemoglobinas variantes na população de Goiás. Para isso a casuística foi composta por 404 alunos participantes dos diversos cursos da Universidade Católica de Goiás (UCG), oriundos de 55 cidades do estado de Goiás. A prevalência de anemia hereditária por talassemias e hemoglobinas variantes em Goiás foi de 10,1%, cuja ordem decrescente foi a seguinte: talassemia alfa heterozigótica (5,2%), heterozigose para hemoglobina S (Hb AS) (2,2%), heterozigose para hemoglobina C (Hb AC) (1%), talassemia beta menor (0,7%), associação entre talassemia alfa e heterozigose para Hb S (0,5%), associação entre talassemia alfa e heterozigose para Hb C (0,3%) e heterozigose para hemoglobina D (Hb AD) (0,3%). Nenhum caso de homozigose foi encontrado no presente estudo. Este trabalho demonstrou a dispersão dos genes para Hb S, Hb C e Hb D, bem como de talassemias alfa e beta em uma população do estado de Goiás. Por essa razão, concluímos que é importante realizar programas com maior abrangência da população para estudo da epidemiologia das talassemias e hemoglobinas variantes no estado de Goiás.

An improved electrophoretic method for a screening program for haemoglobinopathies

A method for a screening program for haemoglobinopathies in a starch agar gel mixed with saponin is presented. Normal and abnormal blood containing haemoglobins S, C, I, M Boston, D Punjab, beta thalassaemia major and beta thalassaemia minor, were applied, in a tray with the capacity for 100 samples. The electrophoresis was performed in 45 min using 300 V. This method offers special advantages for the examination of a large number of samples, using a small amount of whole blood and without the previous preparation of haemoglobin solution.

Diagnóstico laboratorial de hemoglobinopatias em populações diferenciadas

The inherited haemoglobinopathies are a heterogeneous group of recessive disorders that include the thalassaemias and sickle cell disease. Nearly a thousand mutant alleles have now been characterized. The mutations are regionally specific and in most cases the geographical and ethnic distribution shave been determined providing the foundation for a program of control through screening, genetic counseling and prenatal diagnosis. The diagnosis of hemoglobinopathies requires care for the methodologies applied and the population group which will be evaluated. The information about the abnormal hemoglobin, the medical and psychological aspects and genetic counseling of the carriers and their families are goals of great importance for the success of preventive programs in this area. Aiming to evaluate the laboratory methods for hemoglobinopathy screening and their use in clinical laboratories, we have compared abnormal hemoglobins incidence in the different population groups: blood donors, anemia carriers, newborn and students. The laboratory methods applied involved eletrophoretic proceedings, cytological and biochemical analysis. Within the period from September 1999 through January 2000, we analyzed 524 individuals with varied types of abnormal hemoglobins. Among blood donors, we diagnosed two sickle cell carriers, which suggest the necessity for better care in the process of selection of blood donor candidates. The current interest in the medical and social aspects of sickle cell anemia has resulted in a great increase in methodology research leading to the development of sickle cell screening techniques.

Hemoglobin as AS/alfa talassemia - Importância diagnóstica

Sickle Cell disease is a generic term for a group of genetic disorders characterized by the predominance of hemoglobin S. These disorders include Sickle Cell anemia, the Sickle Cell beta Thalassemia syndromes and Hemoglobinopathies in which hemoglobin S is in association with another abnormal hemoglobin, such as hemoglobin S/C. The Sickle Cell trait (hemoglobin AS) associated with Alpha Thalassemia presents alterations in the red blood cells morphology, usually absent in the heterozygous for this hemoglobin variant. The interaction between hemoglobin Sand alpha Thalassemia has been described as one of the factors responsible for the improvement in the clinical picture of homozygous of hemoglobin S (Sickle Cell Anemia), decreasing the number of episodes of pain. The genetic mechanisms of this influence are evaluated using molecular analyses of the human globin genes. With the objective of verifying the presence of alpha Thalassemia in heterozygous of hemoglobin S, with anemia, sent to the Laboratory of Hemoglobins, Department of Biology, UNESP, São José do Rio Preto, SP, we analyzed 1002 blood samples with Sickle Cell trait, in the period from 1990 to 1998. The samples were picked with EDTA 5% as anticoagulant, after previous authorization of the carriers. Appropriated counseling and management requires definitive diagnosis. For the laboratorial diagnosis the blood samples were submitted to electrophoretic procedures in alkaline and acid pH and cytological evaluation of hemoglobin H. The electrophoretic procedures confirmed the presence of hemoglobin AS. The cytological evaluation evidenced the presence of alpha Thalassemia. Of this total analyzed, 16(1,59%) blood samples presented the association between hemoglobin AS and alpha Thalassemia and two individuals belonged of the same family. Our results addressed us to suggest to the routine laboratories, that is important to accomplish the research of alpha Thalassemia among the Sickle Cell trait, with anemia, to verify the interaction with alpha Thalassemia, supplying to the carriers a important information on its hematological profile, genetic pattern of hemoglobinopathies and the appropriated counseling. Rev.bras.hematol.hemoter.,2000,22(3):388-394.

Hemoglobinas anormais e dificuldade diagnóstica

The human hemoglobins, with genetically defined inheritance patterns, have shown characteristic polymorphic variation within the Brazilian population, depending on the racial groups of each region. They have appeared under the form of hemoglobin variants or thalassemias, the variant types S and C and the alpha and beta thalassemias being more common, all of them in heterozygote form. During the year of 1999, blood samples from 506 individuals, with suspected anemia or that had already passed through hemoglobinopathies screening, were sent to the Hemoglobin Reference Center - UNESP for diagnostic confirmation and submitted to electrophoresis proceedings, biochemical and cytological analyses in order to characterize the type of abnormal hemoglobins. The goal of the present study was to verify which abnormal hemoglobin types show greater diagnostic difficulty. The samples came from 24 cities in twelve states. The results showed that 354 (69.96%) individuals presented abnormal hemoglobins, 30 (5.93%) being Hb AS, 5 (0.98%) being Hb AC, 76 (15.02%) suggestive of heterozygote alpha thalassemia, 134 (26.48%) suggestive of heterozygote beta thalassemia and 109 (21.54%) with other forms of abnormal hemoglobin, including rare variants and different forms of thalassemias and variant hemoglobin interactions. It has been concluded that, despite the improved techniques currently available and a constant influx of capacitated personnel, the heterozygote form of thalassemias (210 individuals -41.50%) is challenging to diagnose, followed in difficulty by rare variant characterization and interactive forms of hemoglobinopathies (109 individuals-21,54%), suggesting that the capacity for production of qualified professionals and information about these genetic changes in our population should be increased.

Prevenção de hemoglobinopatias a partir do estudo em gestantes

The Brazilian population, presents genes for abnormal hemoglobins with variable frequencies, which are influenced by the founding racial groups. Thus, the detection of carriers of the genetic alterations is important for public health, since they represent sources of new beterozygotes and possible homozygotes. The control of the hemoglobin pathologies has been possible by means of genetic counseling and early diagnosis. The clinical follow-up of the homozygotes and the orientation of the beterozygotes and especially the couples at risk represent a more effective mode of acting to avoid the birth of children who are carriers of a genetic disease, that is frequently lethal. For these reasons this work had as its objectives: to evaluate the importance of testing in pregnant women for the detection of hemoglobin pathologies with the purpose of investigating the prevalence, attaining prevention, a familial study and awareness; for the positive cases such as couples at risk, orient as to appropriate medical attendance; and to evaluate the response to the program. Of the total of 696 pregnant women analysed, 10.7% revealed hemoglobin pathologies with the following rates: alpha Thalassemia 6.75%; Hb AS 2.01%; beta minor Thalassemia 1.29%; Hb AC 0.28%; Hb AJ 0.14%; Hb AS/Alpha Thalassemia 0.14% and P.H.H.F. 0.14%. The high rates of hemoglobin pathologies encountered in the population of pregnant women studied shows the necessity of the implantation of tests for these abnormalities in the pre-natal routine, since in this period the mothers are more apt to be preoccupied with their own health and that of their babies and, however earlier diagnosed the alterations in the hemoglobins, better and more adequate will be the orientations given the couple.

Utilization of different methodologies for the characterization of Hb Hasharon heterozygotes

Hb Hasharon has an electrophoretic mobility similar to that of Hb S in cellulose acetate and a mobility between Hb S and C at acid pH. In high-performance liquid chromatography, Hb Hasharon shows a distinct chromatographic profile and retention time. The origin of this variant is a mutation in codon 47 (GAC → CAC) of the α2-globin gene, resulting in the replacement of asparagine by histidine during the translation process. Ten blood samples from individuals suspected of being Hb Hasharon carriers were analyzed. In addition to classic laboratory tests and high-performance liquid chromatography, molecular analysis by polymerase chain reaction with restriction fragment length polymorphism designed in the laboratory was performed to confirm this mutation. The study of these cases showed that a combination of classical and molecular methodologies is necessary in the diagnosis of hemoglobinopathies for a correct hemoglobin mutant identification. The accurate identification of hemoglobin variants is essential for genetic counseling and choice of therapy. ©FUNPEC-RP.

HFE gene mutations in Brazilian thalassemic patients

Hereditary hemochromatosis is a disorder of iron metabolism characterized by increased iron intake and progressive storage and is related to mutations in the HFE gene. Interactions between thalassemia and hemochromatosis may further increase iron overload. The ethnic background of the Brazilian population is heterogeneous and studies analyzing the simultaneous presence of HFE and thalassemia-related mutations have not been carried out. The aim of this study was to evaluate the prevalence of the H63D, S65C and C282Y mutations in the HFE gene among 102 individuals with alpha-thalassemia and 168 beta-thalassemia heterozygotes and to compare them with 173 control individuals without hemoglobinopathies. The allelic frequencies found in these three groups were 0.98, 2.38, and 0.29% for the C282Y mutation, 13.72, 13.70, and 9.54% for the H63D mutation, and 0, 0.60, and 0.87% for the S65C mutation, respectively. The chi-square test for multiple independent individuals indicated a significant difference among groups for the C282Y mutation, which was shown to be significant between the beta-thalassemia heterozygote and the control group by the Fisher exact test (P value = 0.009). The higher frequency of inheritance of the C282Y mutation in the HFE gene among beta-thalassemic patients may contribute to worsen the clinical picture of these individuals. In view of the characteristics of the Brazilian population, the present results emphasize the need to screen for HFE mutations in beta-thalassemia carriers.

Three-dimensional visualization of human hemoglobin phenotypes with HPLC

Hemoglobinopathies were included in the Brazilian Neonatal Screening Program on June 6, 2001. Automated high-performance liquid chromatography (HPLC) was indicated as one of the diagnostic methods. The amount of information generated by these systems is immense, and the behavior of groups cannot always be observed in individual analyses. Three-dimensional (3-D) visualization techniques can be applied to extract this information, for extracting patterns, trends or relations from the results stored in databases. We applied the 3-D visualization tool to analyze patterns in the results of hemoglobinopathy based on neonatal diagnosis by HPLC. The laboratory results of 2520 newborn analyses carried out in 2001 and 2002 were used. The Fast, F1, F and A peaks, which were detected by the analytical system, were chosen as attributes for mapping. To establish a behavior pattern, the results were classified into groups according to hemoglobin phenotype: normal (N = 2169), variant (N = 73) and thalassemia (N = 279). 3-D visualization was made with the FastMap DB tool; there were two distribution patterns in the normal group, due to variation in the amplitude of the values obtained by HPLC for the F1 window. It allowed separation of the samples with normal Hb from those with alpha thalassemia, based on a significant difference (P > 0.05) between the mean values of the Fast and A peaks, demonstrating the need for better evaluation of chromatograms; this method could be used to help diagnose alpha thalassemia in newborns.

Planejamento, síntese e avaliação farmacológica de compostos híbridos potencialmente ativos para o tratamento da anemia falciforme

Sickle cell disease (SCD) is a hereditary hemolytic anemia caused by the inheritance of one S hemoglobin gene from each ancestor. Patients with SCD present increased circulating levels of cytokines, including TNF-alpha (TNF-α). Hydroxyurea (HU) is the available therapeutically strategy for treatment; it acts as a source of nitric oxide and benefits patients by increasing the levels of fetal hemoglobin (HbF). Thus, within one research line that aims at finding new drugs, a series of compounds with TNF-α inhibition and nitric oxide donation properties have been synthesized in order to explore possible synergism of actions beneficial in the treatment of the disease. Six compounds were synthesized: five derivatives of organic nitrates and one of sulfonamide. The compounds, (1,3-dioxo-1,3-dihydro-2Hisoindol-2-yl) methyl nitrate (compound I); (1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl) ethyl nitrate (compound II); 3-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl) benzyl nitrate (compound III);4-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-N-hydroxybenzenesulfonamide (compound IV); 4-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl) benzyl nitrate (compound V) and 2-[4-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl) phenyl]ethyl nitrate (compound VI), were synthesized using linear synthetic methodology, with excellent overall yields. All compounds showed anti-inflammatory and analgesic effects with a reduction in 43%-65% of ear edema in mice and a reduction of 25%-42% of writhing induced by acetic acid. All compounds showed comparable reductions in the leukocyte infiltration capacity and ability to generate nitric oxide. The aryl compounds (III, IV and V) presented less mutagenic activity compared to compounds I, II and VI according to the salmonella mutagenicity assay (Ames test). Compounds IV and VI showed activity in K562 culture cells, with increases in gamma globin gene expression to levels higher than with hydroxyurea suggesting a potential to increase fetal hemoglobin. This data set characterizes new potentially useful drug candidates for the treatment of symptoms of sickle cell anemia.