Open reading frame expressed sequences tags (ORESTES) differ from conventional ESTs by providing sequence data from the central protein coding portion of transcripts. We generated a total of 696,745 ORESTES sequences from 24 human tissues and used a subset of the data that correspond to a set of 15,095 full-length mRNAs as a means of assessing the efficiency of the strategy and its potential contribution to the definition of the human transcriptome. We estimate that ORESTES sampled over 80% of all highly and moderately expressed, and between 40% and 50% of rarely expressed, human genes. In our most thoroughly sequenced tissue, the breast, the 130,000 ORESTES generated are derived from transcripts from an estimated 70% of all genes expressed in that tissue, with an equally efficient representation of both highly and poorly expressed genes. In this respect, we find that the capacity of the ORESTES strategy both for gene discovery and shotgun transcript sequence generation significantly exceeds that of conventional ESTs. The distribution of ORESTES is such that many human transcripts are now represented by a scaffold of partial sequences distributed along the length of each gene product. The experimental joining of the scaffold components, by reverse transcription–PCR, represents a direct route to transcript finishing that may represent a useful alternative to full-length cDNA cloning.

Haptoglobin (Hp) is a plasma glycoprotein, the main biological function of which is to bind free hemoglobin (Hb) and prevent the loss of iron and subsequent kidney damage following intravascular hemolysis. Haptoglobin is also a positive acute-phase protein with immunomodulatory properties. In humans, the HP locus is polymorphic, with two codominant alleles (HP1 and HP2) that yield three distinct genotypes/phenotypes (Hp1-1, Hp2-1 and Hp2-2). The corresponding proteins have structural and functional differences that may influence the susceptibility and/or outcome in several diseases. This article summarizes the available data on the structure and functions of Hp and the possible effects of Hp polymorphism in a number of important human disorders.

Transcribed sequences in the human genome can be identified with confidence only by alignment with sequences derived from cDNAs synthesized from naturally occurring mRNAs. We constructed a set of 250,000 cDNAs that represent partial expressed gene sequences and that are biased toward the central coding regions of the resulting transcripts. They are termed ORF expressed sequence tags (ORESTES). The 250,000 ORESTES were assembled into 81,429 contigs. Of these, 1,181 (1.45%) were found to match sequences in chromosome 22 with at least one ORESTES contig for 162 (65.6%) of the 247 known genes, for 67 (44.6%) of the 150 related genes, and for 45 of the 148 (30.4%) EST-predicted genes on this chromosome. Using a set of stringent criteria to validate our sequences, we identified a further 219 previously unannotated transcribed sequences on chromosome 22. Of these, 171 were in fact also defined by EST or full length cDNA sequences available in GenBank but not utilized in the initial annotation of the first human chromosome sequence. Thus despite representing less than 15% of all expressed human sequences in the public databases at the time of the present analysis, ORESTES sequences defined 48 transcribed sequences on chromosome 22 not defined by other sequences. All of the transcribed sequences defined by ORESTES coincided with DNA regions predicted as encoding exons by GENSCAN. (http:͞͞genes.mit.edu/GENSCAN.html). C omplete bacterial genome sequences allow a relatively precise and complete analysis of constituent genes and coding regions by means of direct computational analysis (1). In complex eukaryotic genomes, however, it is proving considerably more difficult to identify genes because of their fragmentation into multiple small exons divided by often considerably larger introns. In this context, the determination of the complete sequence of the human chromosome 22 allowed a detailed appraisal of the efficacy of gene prediction methodologies (2). It was noted that when known genes (where complete cDNA sequences have been determined) were compared with an ab initio prediction of the same region by using the best computational methods available, only 94% of annotated genes were detected. More importantly, in only 20% of cases were all exons exactly predicted, and 16% of all known exons were entirely missed. On the other hand, almost 40% of GENSCAN-predicted genes did not form part of any gene confirmed by other means and include an unknown proportion of false positives (2).In the absence of adequate computational approaches, gene identification will depend on the alignment of finished genomic sequence with sequences from experimentally validated transcripts. Following this approach, Dunham and colleagues (2) were able to identify 247 genes corresponding to fully sequenced transcripts on chromosome 22 that they have denominated Abbreviations: EST, expressed sequence tag; ORESTES, ORF ESTs.cc To whom reprint requests should be addressed.

ResumoObjetivo: Sumarizar os dados disponíveis na literatura recente sobre os aspectos fisiopatológicos, de diagnóstico e tratamento das doenças falciformes e da talassemia β, hemoglobinopatias hereditárias de maior relevância nas populações. Fontes dos dados AbstractObjective: To summarize recently published data on the pathophysiology, diagnosis and treatment of sickle cell diseases and β-Thalassemias, the most relevant hereditary hemoglobinopathies in the global population.Sources: Searches were run on the MEDLINE and SCIELO databases, limited to the period from 2003 to May 2008, using the terms hereditary hemoglobinopathies, sickle cell diseases and β-thalassemia. Two books and two chapters were also included. Summary of the findings:More than 2,000 articles were identified; those providing the most important information and broadest views were selected.Conclusions: Morbidity and mortality rates from sickle cell diseases and β-thalassemia are still very high and represent an important challenge. Increased understanding of pathophysiological aspects has lead to significant improvements in treatment and prevention of these diseases.

We compared the frequencies of the haptoglobin (Hp) genotypes of 775 Brazilian patients with sickle-cell disease divided into the following age groups: 3 months-5 years, 6-10 years, 11-15 years, 16-20 years, and over 20 years. The last group (>20 years) was also compared with a healthy control group and was further divided into subgroups including only subjects aged 21-30 years (V.a and Control.a) and over 30 years (V.b and Control.b). There was no significant difference in the frequencies of the Hp genotypes between the different patient groups or between the patients and controls. However, the Hp2-2 genotype was always less frequent than the Hp1-1 genotype in the patient groups, whereas the opposite was observed in healthy controls. The frequency of Hp2-2 was 25.0% in patients in the 21-30 years age group and fell to 19.5% in those over 30 years. In the controls, the corresponding frequency was around 28%. Although our results do not allow us to conclude that Hp genotypes on their own confer greater or lesser selective advantage on sickle-cell disease patients in the population studied, this polymorphism may, when combined with other genetic and environmental factors, contribute to the clinical diversity observed in this disease.

ObjectiveTo characterize the socioeconomic and demographic aspects of sickle cell disease patients from the state of Rio Grande do Norte (RN), Northeast Brazil, and their adherence to the recommended treatment.MethodsThis cross-sectional descriptive study was performed at referral centers for the treatment of hematological diseases. One hundred and fifty-five unrelated individuals with sickle cell disease who went to these centers for outpatient visits were analyzed. All the patients, or their caregivers, were informed about the research procedures and objectives, and answered a standardized questionnaire.ResultsThe patients were predominantly younger than 12 years old, self-declared as mulatto, lived in small towns fairly distant from the referral center, and had low education and socioeconomic levels. Individuals who were ten or younger were diagnosed at an earlier age. Almost 50% of the patients were taking hydroxyurea, 91.4% reported having received pneumococcal/meningococcal vaccinations and 76.1% received penicillin as antibiotic prophylaxis. However, the majority of them reported having difficulties following the recommendations of the physicians, mainly in respect to attaining the prescribed medications and transportation to the referral centers.ConclusionThese individuals have a vulnerable socioeconomic situation that can lead to an aggravation of their general health and thus deserve special attention from the medical and psychosocial perspectives. Thus, it is necessary to improve public policies that provide Brazilian sickle cell disease patients with better access to medical treatment, living conditions, and integration into society.

Diabetic nephropathy (DN) has an important impact on morbidity/mortality in diabetic patients. Genetic factors are probably involved in the development of this microvascular complication. Haptoglobin (Hp) is a genetically polymorphic glycoprotein that forms stable complexes with plasma-free hemoglobin (Hb) providing protection against heme-induced oxidative stress and kidney damage. The aim of the present study was to investigate the existence of association between the Hp genotypes and the presence of DN in Brazilian diabetic patients. The Hp genotypes of 265 patients, 95 type 1 diabetes mellitus (DM1) sufferers with at least 10 years of disease and 170 type 2 diabetes mellitus (DM2) sufferers with at least 5 years of disease were determined by allele-specific PCR; both groups included patients with and without DN. Hp allele and genotype frequencies were compared among the patient groups and between the patient groups and a control group of 142 healthy individuals. No association between Hp genotypes and DN could be demonstrated. Additionally, urinary albumin excretion values and the presence or absence of systemic arterial hypertension (SAH) were compared among the patient groups. Again, no significant correlations were found. The Hp polymorphism could not be associated with DN in the population studied here.

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