Liane Esteves Daudt

Neonatal screening for hemoglobinopathies: a pilot study in Porto Alegre, Rio Grande do Sul, Brazil

Following a previous evaluation of concept, item and semantic equivalences, this paper assesses the measurement equivalence between a Portuguese version of Revised Conflict Tactics Scales (CTS2) and the original instrument conceived in English. The CTS2 has been widely used to tap violence between couples. An intra-observer reliability evaluation involved 165 replications carried out within a 24-48 hour period. Kappa point-estimates were above 0.75 for all scales except sexual coercion. The analysis of internal consistency concerned 768 subjects with complete sets of items. Kuder-Richardson-20 estimates ranged from 0.65 to 0.86. Results were similar to those found in the original instrument in English for the negotiation, psychological aggression and physical violence scales, yet not so for the sexual coercion and injury scales. Factor analysis identified factors with a recognizable correspondence to the underlying dimensions, although a few inconsistencies were detected. For the assessment of construct validity (n = 528) associations between the instruments scales were evaluated, as well as the relationships between violence and putative underlying dimensions. Overall, the findings suggest that the version can be used in the Brazilian context, although further investigation should be carried out to unveil some important remaining issues.This study was conducted to establish the frequency of hemoglobinopathies among newborns undergoing screening tests for metabolic diseases at the University Hospital (Hospital de Clínicas) in Porto Alegre, Rio Grande do Sul, Brazil. Testing for abnormal hemoglobins was performed by isoelectric focusing electrophoresis on agarose gel with blood obtained by heel stick and applied to filter paper. For confirmatory testing of abnormal neonatal screening, a venopuncture blood sample was obtained from the infant and parents and then submitted to hemoglobin electrophoresis on cellulose acetate at pH 8.6 and citrate agar at pH 6.2. A total of 1,615 subjects were studied: 20 samples showed the Hb S pattern and six samples showed Hb C. Thus, frequency of the sickle cell gene was 1.2% and that of the Hb C gene was 0.4%, regardless of race or origin. These data suggest that the inclusion of universal neonatal screening for hemoglobinopathies in the ongoing projects for the detection of phenylketonuria and congenital hypothyroidism has many advantages and should be considered in health programs.

Triagem neonatal para hemoglobinopatias: um estudo piloto em Porto Alegre, Rio Grande do Sul, Brasil

This study was conducted to establish the frequency of hemoglobinopathies among newborns undergoing screening tests for metabolic diseases at the University Hospital (Hospital de Clinicas) in Porto Alegre, Rio Grande do Sul, Brazil. Testing for abnormal hemoglobins was performed by isoelectric focusing electrophoresis on agarose gel with blood obtained by heel stick and applied to filter paper. For confirmatory testing of abnormal neonatal screening, a venopuncture blood sample was obtained from the infant and parents and then submitted to hemoglobin electrophoresis on cellulose acetate at pH 8.6 and citrate agar at pH 6.2. A total of 1,615 subjects were studied: 20 samples showed the Hb S pattern and six samples showed Hb C. Thus, frequency of the sickle cell gene was 1.2% and that of the Hb C gene was 0.4%, regardless of race or origin. These data suggest that the inclusion of universal neonatal screening for hemoglobinopathies in the ongoing projects for the detection of phenylketonuria and congenital hypothyroidism has many advantages and should be considered in health programs.

Interleukin-15 favors the expansion of central memory CD8+ T cells in ex vivo generated, antileukemia human cytotoxic T lymphocyte lines.

We demonstrated in previous studies that interleukin (IL) -2 supports in vitro cell proliferation of donor-derived cytotoxic T lymphocyte (CTL) lines directed against different types of leukemia blasts. The aim of this study was to compare the capacity of IL-15 with that of IL-2 in supporting the proliferation and cytotoxic activity of antileukemia CTL cultures, and their influence on T-cell memory compartment differentiation. Antileukemia CTL lines were generated using donor-derived dendritic cells pulsed with apoptotic leukemia blasts, in the presence of IL-12 and IL-7, during the primary culture, and expanded through 2 rounds of leukemia-specific stimulation and 1 round of antigen-independent expansion, each supplemented with either IL-2 or IL-15. Both IL-2–supplemented (IL-2–CTLs) and IL-15–supplemented (IL-15–CTLs) lines contained predominant numbers of CD45RA−/CCR7− effector memory (TEM) and CD45RA+/CCR7− (TEMRA+) T cells. Significantly higher numbers (P<0.05) of CD8-positive central memory T cells (TCM), and higher expansion rate, together with comparable cytotoxic activity, were observed in IL-15–CTLs compared with IL-2–CTLs. Altogether, these results demonstrate that IL-15 enhances recovery of CTL activity, without loss of leukemia-directed specificity, and favors expansion of TCM CD8-positive cells, expected to exhibit long-term survival and differentiation capacity in vivo in the presence of a limited amount of antigen.

Single-Cell Cloning of Human, Donor-Derived Antileukemia T-Cell Lines for In vitro Separation of Graft-versus-Leukemia Effect from Graft-versus-Host Reaction

In previous studies, we showed the possibility of expanding in vitro polyclonal CTL lines directed against patient leukemia cells using effector cells derived from both HLA-matched and HLA-mismatched hematopoietic stem cell donors. Some CTL lines, especially those derived from an HLA-disparate donor, displayed residual alloreactivity against patient nonmalignant cells. In this study, we evaluated the possibility of separating in vitro CTLs with selective graft-versus-leukemia (GVL) activity from those potentially involved in the development of graft-versus-host disease (GVHD) through single T-cell cloning of antileukemia polyclonal CTL lines. We showed that CTLs that were expanded from a single T-cell clone (TCC), able to selectively kill leukemia blasts and devoid of alloreactivity towards nonmalignant cells, can be obtained from antileukemia alloreactive polyclonal CTL lines. TCCs expressed a wide repertoire of different T-cell receptor (TCR)-Vβ families, mainly produced IFNγ and interleukin 2, irrespective of CD8 or CD4 phenotype, and could be extensively expanded in vitro without losing their peculiar functional features. The feasibility of our approach for in vitro separation of GVL from GVH reaction opens perspectives for using TCCs, which are selectively reactive towards leukemia blasts, for antileukemia adoptive immune therapy approaches after hematopoietic stem cell transplantation, in particular from HLA-mismatched donors. (Cancer Res 2006; 66(14): 7310-6)

High prevalence of anemia in children and adult women in an urban population in southern Brazil.

This population-based study was designed to detect the prevalence of anemia in a healthy population of children (18 months to 7 years) and women (14 to 30 years) tested in 2006–2007 in the state of Rio Grande do Sul, Brazil as part of an effort to tackle this massive problem that still affects so many people in the XXI century. Anemia was defined according to the WHO. Capillary blood was measured and socioeconomic status was determined according to the Brazilian Association of Market Research Agencies. The median prevalence of anemia in 2198 children was 45.4% and in 1999 women 36.4%. Anemia decreased with age during childhood; although significantly more prevalent in lower classes individuals, it was also high in the upper classes. There are indirect evidences that the lack of iron supplementation and/or iron fortified food may play a role in it. Professionals and society wise measures of education have to be implemented in order to address possible biologic factors involved in childhood psychosocial development in southern Brazil.

Assessment of immature platelet fraction and immature reticulocyte fraction as predictors of engraftment after hematopoietic stem cell transplantation

Engraftment is a critical milestone of the hematopoietic stem cell transplantation (HSCT) process. The immature platelet fraction (IPF) and immature reticulocyte fraction (IRF) are considered early indicators of bone marrow recovery. The objective of this study was to assess these parameters as predictors of HSCT engraftment.

Erratum to “DNMT3A Mutations in Patients with Acute Myeloid Leukemia in South Brazil”

The purpose of this note is to correct both the analysis and typographical errors. The typos are as follows.   The abstract, line 6: In “in 6 patients (8%)” should read “in 5 patients (6%)”.   Page 4, line 12: In “8% (6) of the samples, being 5 missens” should read “6% (5) of the samples, being 4 missense”.   Page 4, line 19: In “Of the 6 cases” should read “Of the 5 cases”.   Page 4, line 20: In “(5, or 83.3%)” should read “(4, or 80%)”.   Page 4, second column, line 8: In “OS for patients with wild DNMT3A gene was 41.4% and for patients with mutated DNMT3A was 44.4% (P = 0.59)” should read “OS for patients with wild DNMT3A gene was 45.7% and for patients with mutated DNMT3A was 60.0% (P = 0.47)”.   Page 6, line 4: In “found in 8%” should read “found in 6%”.   Page 6, second column, line 13: In “3 of 6 mutations” should read “3 of 5 mutations”.   Page 6, second column, line 16: In “Five, or 80%,” should read “Four, or 80%,”.   Page 6, second column, line 18: In “(P = 0.28)” should read “(P = 0.34)”.   Page 6, second column, line 20: In “(20.7 × 109/L)” should read “(15.6 × 109/L)”.   Table 4, column 2, line 6: In “H896*” should read “P896*”. Table 4 Description of somatic mutations found in gene DNMT3A.   Table 4: Entire line 7 was excluded.   Table 5, line 2: In “40.2, 44.8 and 0.56” should read “47.4, 40.4 and 0.42”. Table 5 Clinical characteristics of patients with Acute Myeloid Leukemia with or without DNMT3A mutations.   Table 5, line 4: In “50% (3), 59.3% (45) and 0.68” should read “60% (3), 57.9% (44) and 0.92”.   Table 5, line 5: In “50% (3), 40.7% (31)” should read “40% (2), 42.1% (32)”.   Table 5, line 19: In “50% (3), 51.4% (37) and 1.000” should read “40% (2), 52.7% (39) and 0.66”.   Table 5, line 20: In “50% (2), 30.9% (17) and 0.58” should read “40% (2), 29.2% (21) and 0.63”.   We changed Figure 3 as shown above. Figure 3

Outcome of Treatment in Adult Acute Lymphoblastic Leukemia in Southern Brazil Using a Modified German Multicenter Acute Lymphoblastic Leukemia Protocol

Reports on treatment outcomes in adults with acute lymphoblastic leukemia (ALL) in Brazil are sparse. To evaluate the outcome of patients with ALL managed by the public healthcare system, we studied 42 adults treated from 1990 to 1997 in the Division of Hematology at Hospital de Clínicas, Porto Alegre, Brazil. Of these patients, 14/42 were females and their median age at diagnosis was 26 (17–64) years. The diagnosis of ALL was based on cytological examination of marrow smears, and immunophenotypic and cytogenetic studies, when available. Fifty percent of the patients expressed CD10, 30% were CD10 negative and CD19 positive and 20% expressed T markers. Philadelphia chromosome was found in 4 (7.14%). The chemotherapy protocol was adapted from the German Multicenter ALL (GMALL) 02-84 protocol. The complete remission rate was 93% and the overall survival at 5 years was 41%. No particular risk factor was identified in our series. These results are comparable to the findings of other international studies.

DNMT3A Mutations in Patients with Acute Myeloid Leukemia in South Brazil

Acute myeloid leukemia (AML) is a complex and heterogeneous hematopoietic tissue neoplasm. Several molecular markers have been described that help to classify AML patients into risk groups. DNA methyltransferase 3A (DNMT3A) gene mutations have been recently identified in about 22% of AML patients and associated with poor prognosis as an independent risk factor. Our aims were to determine the frequency of somatic mutations in the gene DNMT3A and major chromosomal translocations in a sample of patients with AML. We investigated in 82 samples of bone marrow from patients with AML for somatic mutations in DNMT3A gene by sequencing and sought major fusion transcripts by RT-PCR. We found mutations in the DNMT3A gene in 6 patients (8%); 3 were type R882H. We found fusion transcripts in 19 patients, namely, AML1/ETO (n = 5; 6.1%), PML/RARα (n = 12; 14.6%), MLL/AF9 (0; 0%), and CBFβ/MYH11 (n = 2; 2.4%). The identification of recurrent mutations in the DNMT3A gene and their possible prognostic implications can be a valuable tool for making treatment decisions. This is the first study on the presence of somatic mutations of the DNMT3A gene in patients with AML in Brazil. The frequency of these mutations suggests a possible ethnogeographic variation.

The role of BCL11A and HMIP-2 polymorphisms on endogenous and hydroxyurea induced levels of fetal hemoglobin in sickle cell anemia patients from southern Brazil.

High levels of fetal hemoglobin (HbF) reduce sickle cell anemia (SCA) morbidity and mortality. HbF levels vary considerably and there is a strong genetic component that influences HbF production. Genetic polymorphisms at three quantitative trait loci (QTL): Xmn1-HBG2, HMIP-2 and BCL11A, have been shown to influence HbF levels and disease severity in SCA. Hydroxyurea (HU) is a drug that increases HbF. We investigated the influence of single nucleotide polymorphisms (SNPs) at the Xmn1-HBG2 (rs7482144); BCL11A (rs1427407, rs4671393 and rs11886868); and HMIP-2 (rs9399137 and rs9402686) loci on baseline and HU-induced HbF levels in 111 HbSS patients. We found that both BCL11A and HMIP-2 were associated with increased endogenous levels of HbF. Interestingly, we also found that BCL11A was associated with higher induction of HbF with HU. This effect was independent of the effect of BCL11A on baseline HbF levels. Additional studies will be needed to validate these findings and explain the ample inter-individual variations in HbF levels at baseline and HU-induced in patients with SCA.