Evaluation of computational methods for the reconstruction of HLA haplotypes

Human leukocyte antigen (HLA) haplotypes are frequently evaluated for population history inferences and association studies. However, the available typing techniques for the main HLA loci usually do not allow the determination of the allele phase and the constitution of a haplotype, which may be obtained by a very time-consuming and expensive family-based segregation study. Without the family-based study, computational inference by probabilistic models is necessary to obtain haplotypes. Several authors have used the expectation-maximization (EM) algorithm to determine HLA haplotypes, but high levels of erroneous inferences are expected because of the genetic distance among the main HLA loci and the presence of several recombination hotspots. In order to evaluate the efficiency of computational inference methods, 763 unrelated individuals stratified into three different datasets had their haplotypes manually defined in a family-based study of HLA-A, -B, -DRB1 and -DQB1 segregation, and these haplotypes were compared with the data obtained by the following three methods: the Expectation-Maximization (EM) and Excoffier-Laval-Balding (ELB) algorithms using the arlequin 3.11 software, and the PHASE method. When comparing the methods, we observed that all algorithms showed a poor performance for haplotype reconstruction with distant loci, estimating incorrect haplotypes for 38%-57% of the samples considering all algorithms and datasets. We suggest that computational haplotype inferences involving low-resolution HLA-A, HLA-B, HLA-DRB1 and HLA-DQB1 haplotypes should be considered with caution.

Low-Carbohydrate and High-Fat Diets on the Promotion of Hepatic Steatosis in Rats

Aim: The present work looked for to evaluate in rats the impact of different diets (high-lipid and high-lipid + high-protein) on liver, verifying the occurrence of oxidative stress and steatosis. Methods: The animals were treated with the respective diets (Group HLS: high-lipid diet with 50% of saturated fat; Group HPLS: high-lipid and high-protein diet with 50% of saturated fat and 40% of protein; Group Control: control diet AIN-93) for 28 days. After this period the animals were sacrificed for hepatic determinations of MDA, reduced GSH, vitamin E, steatosis and glycemia. Results: The results showed higher glycemia in the group HPLS, high concentration of MDA and GSH in the group Control and decreased hepatic vitamin E concentration in the groups that received the high-lipid diets. The hepatic fat was higher in the groups HPLS and HLS in relation to the Group Control, however HPLS presenting high level of fat concentration, showing similar results as the steatosis. Conclusion: the fat increase in the diet promoted increase of the oxidative stress, evidenced by the decrease in the hepatic concentration of vitamin E, showing its antioxidant role against the probable generated free radicals, the ones which possibly exercised a role in the steatosis occurrence.

Under-reporting of food intake is frequent among Brazilian free-living older persons: a doubly labelled water study

The assessment of food intake is essential for the development of dietetic interventions. Accuracy is low when intake is assessed by questionnaires, the under-reporting of food intake being frequent. Most such studies, however, were performed in developed countries and there is little data about the older population of developing nations. This study aimed to verify the total energy expenditure (TEE) of independent older Brazilians living in an urban area, through the doubly labelled water (DLW) method and to compare it with the reported energy intake obtained through the application of a food frequency questionnaire (FFQ). Initially, 100 volunteers aged from 60 to 75 years had their body composition determined by dual-energy X-ray absorptiometry (DEXA). Five volunteers of each quartile of body fat percentage had their energy expenditure determined by DLW. The mean age of the subjects included in this phase of the study was 66.4 +/- 3.5 years, and ten of the subjects were men. The mean TEE was 2565 +/- 614 and 2154 +/- 339 kcal.day(-1) for men and women, respectively. The Physical Activity Level (PAL) was 1.58 +/- 0.31 and 1.52 +/- 0.22, respectively. Under-reporting of food intake was highly prevalent, with a mean percentage of reported intake in relation to measured TEE of -17.7%. Thus, under-reporting of food intake is highly prevalent among Brazilian independent older persons. The DLW method is an important tool in nutritional studies and its use is to be recommended in developing countries. Copyright (C) 2010 John Wiley & Sons, Ltd.

Comparative analysis of two immunohistochemical methods for antigen retrieval in the optical lobe of the honeybee Apis mellifera: Myosin-v assay

The present study compared two heating methods currently used for antigen retrieval (AR) immunostaining: the microwave oven and the steam cooker. Myosin-V, a molecular motor involved in vesicle transport, was used as a neuronal marker in honeybee Apis mellifera brains fixed in formalin. Overall, the steam cooker showed the most satisfactory AR results. At 100 degrees C, tissue morphology was maintained and revealed epitope recovery, while evaporation of the AR solution was markedly reduced; this is important for stabilizing the sodium citrate molarity of the AR buffer and reducing background effects. Standardization of heat-mediated AR of formalin-fixed and paraffin-embedded tissue sections results in more reliable immunostaining of the honeybee brain.

Methods for exploring the morpho-functional relations of the aortic depressor nerve in experimental diabetes

The present study investigated morpho-functional relations of the aortic depressor nerve (ADN) 5, 15 and 120 days after the onset of streptozotocin-induced diabetes in rats. Time control animals received vehicle. Under pentobarbital anesthesia, ADN activity was recorded simultaneously with arterial pressure. After the recordings, nerves were prepared for light microscopy study and morphometry. ADN function was accessed by means of pressure-nerve activity curve (fitted by sigmoidal regression) and cross-spectral analysis between mean arterial pressure (MAP) and ADN activity. The relation between morphological (myelinated fibers number and density, total myelin area, total fiber area and percentage of occupancy) and functional (gain, signal/noise relation, frequency) parameters were accessed by linear regression analysis and correlation coefficient calculations. Functional parameters obtained by means of the sigmoidal regression curve as well as by cross-spectral analysis were similar in diabetic and control rats. Morphometric parameters of the ADN were similar between groups 5 days after the onset of diabetes. Average myelin area and myelinated fiber area were significantly smaller on diabetic rats 15 and 120 days after the onset of diabetes, being the myelinated fiber and respective axons area and diameter also smaller on 120 days group. Nevertheless, G ratio (ratio between axon and fiber diameter) was nearly 0.6 and not different between groups or experimental times. No significant relationship between morphological and functional parameters was detected in all experimental groups. The present study suggests that ADN diabetic neuropathy was time-dependent, with damage to myelinated fibers to be the primary event, not evidenced by physiological methods. (C) 2010 Elsevier B.V. All rights reserved.

Increased circulating cell-free hemoglobin levels reduce nitric oxide bioavailability in preeclampsia

Contrasting with increased nitric oxide (NO) formation during healthy pregnancy, reduced NO bioavailability plays a role in preeclampsia. However, no study has examined whether increased NO consumption by enhanced circulating levels of cell-free hemoglobin plays a role in preeclampsia. We studied 82 pregnant women (38 healthy pregnant and 44 with preeclampsia). To assess NO bioavailability, we measured plasma and whole blood nitrite concentrations using an ozone-based chemiluminescence assay. Plasma ceruloplasmin concentrations and plasma NO consumption (pNOc) were assessed and plasma hemoglobin (pHb) concentrations were measured with a commercial immunoassay. We found lower whole blood and plasma nitrite concentrations in preeclamptic patients (-48 and -39%, respectively; both P<0.05) compared with healthy pregnant women. Plasma samples from preeclamptic women consumed 63% more NO (P=0.003) and had 53% higher pHb and 10% higher ceruloplasmin levels than those found in healthy pregnant women (P<0.01). We found significant positive correlations between pHb and pNOc (r=0.61; P<0.0001), negative correlations between pNOc and whole blood or plasma nitrite concentrations (P=0.02; r=-0.32 and P=0.01: r=-0.34, respectively), and negative correlations between pHb and whole blood or plasma nitrite concentrations (P=0.03; r=-0.36 and P=0.01: r=-0.38, respectively). These findings suggest that increased pHb levels lead to increased NO consumption and lower NO bioavailability in preeclamptic compared with healthy pregnant women. (C) 2010 Elsevier Inc. All rights reserved.

Impact of Surgical Staging in Locally Advanced Cervical Cancer and Subsequent Chemotherapy

Background: Surgical staging (SS) is the gold standard for determination of the true extent of a patient`s disease and is an important prognostic factor in cervical cancer. We investigated whether lymph node dissection (LND) prior to chemotherapy (CT) followed by radical surgery (RS) could modified overall (OS) and disease-free survival (DFS). Methods: We performed a cohort analysis of 98 patients with cervical carcinoma. The experimental group consisted of 36 patients who underwent SS followed by neoadjuvant chemotherapy, and then by RS (objective response) or chemo-radiation therapy (with or without subsequent surgery when not possible). The control group consisted of 62 similarly treated patients without pretreatment SS. The value of this procedure as a diagnostic tool in defining the extent of disease was evaluated. Furthermore, LND/CT-associated treatment complications and the impacts on OS and DFS were also evaluated. Results: Fourteen (38.9%) patients had pelvic LN metastases and three (8.3%) patients had pelvic and para-aortic LN metastases. The 39-month OS and DFS rates for the current study were 90.6% for the staged group and 52% for non-staged treatment (P < 0.001). Conclusion: SS in cervical cancer is a feasible and safe pretreatment procedure, and when associated with CT, it improves OS and DFS. J. Surg. Oncol. 2009;100:505-510. (C) 2009 Wiley-Liss, Inc.

Morphological comparison of different protocols of skeletal muscle remobilization in rats after hindlimb suspension

The aim of this study was to evaluate and compare the efficacy of different remobilization protocols in different skeletal muscles considering the changes induced by hindlimb suspension of the tail. Thirty-six female Wistar rats were divided into six groups: control I, control II, suspended, suspended free, suspended trained on a declined treadmill and suspended trained on a flat treadmill. Fragments of soleus and tibialis anterior (TA) muscle were frozen and processed by different histochemical methods. The suspended soleus showed a significant increase in the proportional number of intermediate/hybrid fibers and a decrease in the number of type I fibers. Some of these changes proved to be reversible after remobilization. The three remobilization programs led to the recovery of both the proportional number of fibers and their size. The TA muscle presented a significant increase in the number and size of type I fibers and a cell size reduction of type IIB fibers, which were recovered after training on a declined treadmill and free movement. Especially regarding the soleus, the present findings indicate that, among the protocols, training on a declined treadmill was found to induce changes of a more regenerative nature, seemingly indicating a better tissue restructuring after the suspension procedure.

Efficacy of cidofovir in recurrent juvenile respiratory papillomatosis

The efficacy of cidofovir in juvenile recurrent respiratory papillomatosis (JRRP) remains uncertain due to the lack of published case-control studies. Aim: To establish factors affecting the progression of JRRP prognosis, and to evaluate cidofovir for eradicating JRRP. Study Design: Retrospective. Methods: 22 children with JRRP were evaluated at a referral center. All children underwent surgical debulking, followed by cidofovir injection (Group 2) or not (Group 1). Age at diagnosis was correlated with the Derkay score and disease outcome. Disease progression was compared between groups 1 and 2. Results: fifteen children were considered disease-free; 8 were in Group 2 and 7 in Group 1. Age and total and clinical scores (P<0.05) were negatively correlated. The mean number of surgeries required to control the disease was identical in both groups; the duration of treatment until remission was significantly higher in Group 1 (P<0,05). Conclusion: JRRP is more aggressive in earlier onset disease. The duration of treatment was significantly lower in patients treated with cidofovir until eradication of JRRP compared to patients treated with surgery only.

Vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (NOS3) polymorphisms are associated with high relapse risk in childhood acute lymphoblastic leukemia (ALL)

Background: Angiogenesis has been shown as an important process in hematological malignancies. It consists in endothelial proliferation, migration, and tube formation following pro-angiogenic factors releasing, specially the vascular endothelial growth factor (VEGF), which angiogenic effect seems to be dependent on nitric oxide (NO). We examined the association among functional polymorphism in these two angiogenesis related genes: VEGF (-2578C>A, -1154G>A, and -634G>C) and NOS3 (-786T>C, intron 4 b>a, and Glu298Asp) with prognosis of childhood acute lymphoblastic leukemia (ALL). Methods: The genotypes were determined and haplotypes estimated in 105 ALL patients that were divided in 2 groups: high risk (HR) and low risk of relapse (LR) patients. In addition, event-free survival curves according to genotypes were assessed. Results: The group HR compared to the LR showed a higher frequency of the alleles -2578C and -634C and the haplotype CGC for VEGF (0.72 vs. 0.51, p<0.008; 0.47 vs. 0.26, p<0.008; and 42.1 vs. 14.5, p<0.006; respectively) and a lower frequency of the haplotype CbGlu (0.4 vs. 8.8, p<0.006), for NOS3. Conclusion: Polymorphisms of VEGF and NOS3 genes are associated with high risk of relapse, therefore may have a prognostic impact in childhood ALL. (C) 2010 Elsevier B.V. All rights reserved.

Benefits of the Intermittent Use of 6-Mercaptopurine and Methotrexate in Maintenance Treatment for Low-Risk Acute Lymphoblastic Leukemia in Children: Randomized Trial From the Brazilian Childhood Cooperative Group Protocol ALL-99

Purpose To describe event-free survival (EFS) and toxicities in children with low-risk acute lymphoblastic leukemia (ALL) assigned to receive either continuous 6-mercaptopurine (6-MP) and weekly methotrexate (MIX) or intermittent 6-MP with intermediate-dose MTX, as maintenance treatment. Patients and Methods Between October 1, 2000, and December 31, 2007, 635 patients with low-risk ALL were enrolled onto Brazilian Childhood Cooperative Group for ALL Treatment (GBTLI) ALL-99 protocol. Eligible children (n=544) were randomly allocated to receive either continuous 6-ME/MIX (group 1, n 272) or intermittent 6-MP (100 mg/m(2)/d for 10 days, with 11 days resting) and MIX (200 mg/m(2) every 3 weeks; group 2, n = 272). Results The 5-year overall survival (OS) and EFS were 92.5% +/- 1.5% SE and 83.6% +/- 2.1% SE, respectively. According to maintenance regimen, the OS was 91.4% +/- 2.2% SE (group 1) and 93.6% +/- 2.1% SE (group 2; P=.28) and EFS 80.9% +/- 3.2% SE (group 1) and 86.5% +/- 2.8% SE (group 2; P=.089). Remarkably, the intermittent regimen led to significantly higher EFS among boys (85.7% v 74.9% SE; P=027), while no difference was seen for girls (87.0% v 88.8% SE; P=.78). Toxic episodes were recorded in 226 and 237 children, respectively. Grade 3 to 4 toxic events for groups 1 and 2 were, respectively, 273 and 166 for hepatic dysfunction (P=.002), and 772 and 636 for hematologic episodes (P=.005). Deaths on maintenance were: seven (group 1) and one (group 2). Conclusion The intermittent use of 6-MP and MIX in maintenance is a less toxic regimen, with a trend toward better long-term EFS. Boys treated with the intermittent schedule had significantly better EFS.

Advances in Imaging of Osteoarthritis and Cartilage

Osteoarthritis (OA) is the most frequent form of arthritis, with major implications for individual and public health care without effective treatment available. The field of joint imaging, and particularly magnetic resonance (MR) imaging, has evolved rapidly owing to technical advances and the application of these to the field of clinical research. Cartilage imaging certainly is at the forefront of these developments. In this review, the different aspects of OA imaging and cartilage assessment, with an emphasis on recent advances, will be presented. The current role of radiography, including advances in the technology for joint space width assessment, will be discussed. The development of various MR imaging techniques capable of facilitating assessment of cartilage morphology and the methods for evaluating the biochemical composition of cartilage will be presented. Advances in quantitative morphologic cartilage assessment and semiquantitative whole-organ assessment will be reviewed. Although MR imaging is the most important modality in imaging of OA and cartilage, others such as ultrasonography play a complementary role that will be discussed briefly.