Induced spawning of the endangered Neotropical species Steindachneridion parahybae (Siluriformes: Pimelodidae)

The "surubim do Paraíba" (Steindachneridion parahybae) is a freshwater catfish endemic to the Paraíba do Sul River basin, Brazil. This species has been seriously threatened by environmental disturbances in the last several decades. Wild Steindachneridion parahybae males and females were collected in 2003 and taken to the hatchery of a power plant of the Companhia Energética de São Paulo (CESP). Steindachneridion parahybae broodstocks were artificially induced to reproduce in December 2003 using a combination of carp pituitary extract (CPE) and human chorionic gonadotropin (hCG). Oocytes and milt were stripped; the fertilized eggs were transferred to 60-liter conical incubators and hatched larvae distributed in nine horizontal trays. Exogenous feed was started just after yolk sac absorption. A high rate of cannibalism and photophobia were observed during the larval period, resulting in a 26% survival rate from larvae to fingerlings.

High Prevalence of Side Effects After Recombinant Human Thyrotropin-Stimulated Radioiodine Treatment with 30 mCi in Patients with Multinodular Goiter and Subclinical/Clinical Hyperthyroidism

Background: Treatment of multinodular goiters (MNGs) is highly controversial. Radioiodine (RAI) therapy is a nonsurgical alternative for the elderly who decline surgery. Recently, recombinant human thyrotropin (rhTSH) has been used to augment RAI uptake and distribution. In this study, we determined the outcome of 30 mCi RAI preceded by rhTSH (0.1 mg) in euthyroid (EU) and hyperthyroid (subclinical/clinical) patients with large MNGs. Methods: This was a prospective cohort study. Forty-two patients (age, 43-80 years) with MNGs were treated with 30 mCi RAI after stimulation with 0.1 mg of rhTSH. Patients were divided into three groups, according to thyroid function: EU (n = 18), subclinically hyperthyroid (SC-H, n = 18), and clinically hyperthyroid (C-H, n = 6). All patients underwent a 90-day low-iodine diet before treatment, and those with clinical hyperthyroidism received methimazole 10 mg daily for 30 days. Serum TSH, free thyroxine (FT4), total triiodothyronine (TT3), and thyroglobulin were measured at baseline and at 24, 48, 72, 168 hours, and 1, 3, 6, 9, 12, 18, 24, and 36 months after therapy. Thyroid volume was assessed by computed tomography at baseline and every 6 months. Results: Patients had high iodine urinary excretion (308 +/- 108 mu g I/L) at baseline. TSH levels at baseline were within the normal range (1.5 +/- 0.7 mu U/mL) in the EU group and suppressed (< 0.3 mu U/mL) in the SC-H and C-H groups. After rhTSH, serum TSH peaked at 24 hours reaching 12.4 +/- 5.85 mu U/mL. After RAI administration, patients in both hyperthyroid groups had a higher increase in FT4 and TT3 compared with those in the EU group (p < 0.001). Thyroglobulin levels increased equally in all three groups until day 7. Thyroid volume decreased significantly in all patients. Side effects were more common in the SC-H and C-H groups (31.4% and 60.4%, respectively) compared with EU patients (17.8%). Permanent hypothyroidism was more prevalent in the EU group (50%) compared with the SC-H (11%) and C-H (16.6%) groups. Conclusions: Patients with MNG may have subclinical and clinical nonautoimmune iodine-induced hyperthyroidism. Despite a low-iodine diet and therapy with methimazole, hyperthyroid patients have a significantly higher increase in FT4 and TT3 levels after RAI ablation. This can lead to important side effects related mostly to the cardiac system. We strongly advise that patients with SC-H and C-H be adequately treated with methimazole and low-iodine diet aiming to normalize their hyperthyroid condition before rhTSH-stimulated treatment with RAI.

Poly (A)(+) Transcriptome Assessment of ERBB2-Induced Alterations in Breast Cell Lines

We report the first quantitative and qualitative analysis of the poly (A)(+) transcriptome of two human mammary cell lines, differentially expressing (human epidermal growth factor receptor) an oncogene over-expressed in approximately 25% of human breast tumors. Full-length cDNA populations from the two cell lines were digested enzymatically, individually tagged according to a customized method for library construction, and simultaneously sequenced by the use of the Titanium 454-Roche-platform. Comprehensive bioinformatics analysis followed by experimental validation confirmed novel genes, splicing variants, single nucleotide polymorphisms, and gene fusions indicated by RNA-seq data from both samples. Moreover, comparative analysis showed enrichment in alternative events, especially in the exon usage category, in ERBB2 over-expressing cells, data indicating regulation of alternative splicing mediated by the oncogene. Alterations in expression levels of genes, such as LOX, ATP5L, GALNT3, and MME revealed by large-scale sequencing were confirmed between cell lines as well as in tumor specimens with different ERBB2 backgrounds. This approach was shown to be suitable for structural, quantitative, and qualitative assessment of complex transcriptomes and revealed new events mediated by ERBB2 overexpression, in addition to potential molecular targets for breast cancer that are driven by this oncogene.

Role of upper endoscopy in diagnosing opportunistic infections in human immunodeficiency virus-infected patients

Highly active antiretroviral therapy (HAART) has dramatically decreased opportunistic infections (OIs) in human immunodeficiency virus (HIV)-infected patients. However, gastrointestinal disease continues to account for a high proportion of presenting symptoms in these patients. Gastrointestinal symptoms in treated patients who respond to therapy are more likely to the result of drug-induced complications than OI. Endoscopi evaluation of the gastrointestinal tract remains a cornerstone of diagnosis, especially in patients with advanced immunodeficiency, who are at risk for OI. The peripheral blood CD4 lymphocyte count helps to predict the risk of an OI, with the highest risk seen in HIV-infected patients with low CD4 count (< 200 cells/mm(3)). This review provides an update of the role of endoscopy in diagnosing OI in the upper gastrointestinal tract in HIV-infected patients in the era of HAART. (C) 2009 The WJG Press and Baishideng. All rights reserved.

Fluorescence and reflectance spectroscopy for identification of atherosclerosis in human carotid arteries using principal components analysis

Objectives: The aim of this work was to verify the differentiation between normal and pathological human carotid artery tissues by using fluorescence and reflectance spectroscopy in the 400- to 700-nm range and the spectral characterization by means of principal components analysis. Background Data: Atherosclerosis is the most common and serious pathology of the cardiovascular system. Principal components represent the main spectral characteristics that occur within the spectral data and could be used for tissue classification. Materials and Methods: Sixty postmortem carotid artery fragments (26 non-atherosclerotic and 34 atherosclerotic with non-calcified plaques) were studied. The excitation radiation consisted of a 488-nm argon laser. Two 600-mu m core optical fibers were used, one for excitation and one to collect the fluorescence radiation from the samples. The reflectance system was composed of a halogen lamp coupled to an excitation fiber positioned in one of the ports of an integrating sphere that delivered 5 mW to the sample. The photo-reflectance signal was coupled to a 1/4-m spectrograph via an optical fiber. Euclidean distance was then used to classify each principal component score into one of two classes, normal and atherosclerotic tissue, for both fluorescence and reflectance. Results: The principal components analysis allowed classification of the samples with 81% sensitivity and 88% specificity for fluorescence, and 81% sensitivity and 91% specificity for reflectance. Conclusions: Our results showed that principal components analysis could be applied to differentiate between normal and atherosclerotic tissue with high sensitivity and specificity.

Detection of Human Bocavirus mRNA in Respiratory Secretions Correlates with High Viral Load and Concurrent Diarrhea

Human bocavirus (HBoV) is a parvovirus recently identified in association with acute respiratory infections (ARI). Despite its worldwide occurrence, little is known on the pathogenesis of HBoV infections. In addition, few systematic studies of HBoV in ARI have been conducted in Latin America. Therefore, in order to test whether active viral replication of human bocavirus is associated with respiratory diseases and to understand the clinical impact of this virus in patients with these diseases, we performed a 3-year retrospective hospital-based study of HBoV in outpatients and inpatients with symptoms of Acute Respiratory Infections (ARI) in Brazil. Nasopharyngeal aspirates (NPAs) from 1015 patients with respiratory symptoms were tested for HBoV DNA by PCR. All samples positive for HBoV were tested by PCR for all other respiratory viruses, had HBoV viral loads determined by quantitative real time PCR and, when possible, were tested by RT-PCR for HBoV VP1 mRNA, as evidence of active viral replication. HBoV was detected in 4.8% of patients, with annual rates of 10.0%, 3.0% and 3.0% in 2005, 2006 and 2007, respectively. The range of respiratory symptoms was similar between HBoV-positive and HBoV-negative ARI patients. However, a higher rate of diarrhea was observed in HBoV-positive patients. High HBoV viral loads (> 10(8) copies/mL) and diarrhea were significantly more frequent in patients with exclusive infection by HBoV and in patients with detection of HBoV VP1 mRNA than in patients with viral co-infection, detected in 72.9% of patients with HBoV. In summary, our data demonstrated that active HBoV replication was detected in a small percentage of patients with ARI and was correlated with concurrent diarrhea and lack of other viral co-infections.

RNA interference-mediated knockdown of CD49e (alpha 5 integrin chain) in human thymic epithelial cells modulates the expression of multiple genes and decreases thymocyte adhesion

Background: The thymus is a central lymphoid organ, in which bone marrow-derived T cell precursors undergo a complex process of maturation. Developing thymocytes interact with thymic microenvironment in a defined spatial order. A component of thymic microenvironment, the thymic epithelial cells, is crucial for the maturation of T-lymphocytes through cell-cell contact, cell matrix interactions and secretory of cytokines/chemokines. There is evidence that extracellular matrix molecules play a fundamental role in guiding differentiating thymocytes in both cortical and medullary regions of the thymic lobules. The interaction between the integrin alpha 5 beta 1 (CD49e/CD29; VLA-5) and fibronectin is relevant for thymocyte adhesion and migration within the thymic tissue. Our previous results have shown that adhesion of thymocytes to cultured TEC line is enhanced in the presence of fibronectin, and can be blocked with anti-VLA-5 antibody. Results: Herein, we studied the role of CD49e expressed by the human thymic epithelium. For this purpose we knocked down the CD49e by means of RNA interference. This procedure resulted in the modulation of more than 100 genes, some of them coding for other proteins also involved in adhesion of thymocytes; others related to signaling pathways triggered after integrin activation, or even involved in the control of F-actin stress fiber formation. Functionally, we demonstrated that disruption of VLA-5 in human TEC by CD49e-siRNA-induced gene knockdown decreased the ability of TEC to promote thymocyte adhesion. Such a decrease comprised all CD4/CD8-defined thymocyte subsets. Conclusion: Conceptually, our findings unravel the complexity of gene regulation, as regards key genes involved in the heterocellular cell adhesion between developing thymocytes and the major component of the thymic microenvironment, an interaction that is a mandatory event for proper intrathymic T cell differentiation.

Evaluation of Protective Effect of a Water-In-Oil Microemulsion Incorporating Quercetin Against UVB-Induced Damage in Hairless Mice Skin

Purpose. Histological aspects were considered in order to evaluate the in vivo photoprotective effect of a w/o microemulsion containing quercetin against UVB irradiation-induced dermal damages. The toxicity in cell culture and the potential skin irritation resulting from topical application of this formulation were investigated. Methods. Mouse dorsal surfaces were treated topically with 300 mg of the unloaded and quercetin-loaded (0.3%, w/w) microemulsions before and after exposure to UVB (2.87 J/cm(2)) irradiation. The untreated control groups irradiated and non-irradiated were also evaluated. UVB-induced histopathological changes as well as the photoprotective effect of this formulation were evaluated considering the parameters of infiltration of inflammatory cells, epidermis thickening (basale and spinosum layers) and collagen and elastic fiber contents. The cytotoxicity of the reported formulation was evaluated in L929 mice fibroblasts by MTT assay and the skin irritation was investigated after topical application of both unloaded and quercetin-loaded microemulsions once a day for 15 days. Results. The results demonstrated that the w/o microemulsion containing quercetin reduced the incidence of histological skin alterations, mainly the connective-tissue damage, induced by exposure to UVB irradiation. This suggests that protective effects of this formulation against UV-induced responses are not secondary to the interference of UV transmission (i.e., blocking the UVB radiation from being absorbed by the skin), as is usually implied with UVB absorbers and sunscreens, but is instead due to different biological effects of this flavonoid. Furthermore, by evaluating the cytotoxic effect on L929 cells and histological aspects such as infiltration of inflammatory cells and epidermis thickness of hairless mice, the present study also demonstrated the lack of toxicity of the proposed system. Conclusion. Based on these mice models, a detailed characterization of the w/o microemulsion incorporating quercetin effects as a photochemoprotective agent on human skin is presented.

The Thermal Effects of Therapeutic Lasers with 810 and 904 nm Wavelengths on Human Skin

Objective: To investigate the effect of therapeutic infrared class 3B laser irradiation on skin temperature in healthy participants of differing skin color, age, and gender. Background: Little is known about the potential thermal effects of Low Level Laser Therapy (LLLT) irradiation on human skin. Methods: Skin temperature was measured in 40 healthy volunteers with a thermographic camera at laser irradiated and control (non-irradiated) areas on the skin. Six irradiation doses (2-12 J) were delivered from a 200mW, 810nm laser and a 60mW, 904nm laser, respectively. Results: Thermal effects of therapeutic LLLT using doses recommended in the World Association for Laser Therapy (WALT) guidelines were insignificant; below 1.5 degrees C in light, medium, and dark skin. When higher irradiation doses were used, the 60mW, 904 nm laser produced significantly (p < 0.01) higher temperatures in dark skin (5.7, SD +/- 1.8 degrees C at 12 J) than in light skin, although no participants requested termination of LLLT. However, irradiation with a 200mW, 810nm laser induced three to six times more heat in dark skin than in the other skin color groups. Eight of 13 participants with dark skin asked for LLLT to be stopped because of uncomfortable heating. The maximal increase in skin temperature was 22.3 degrees C. Conclusions: The thermal effects of LLLT at doses recommended by WALT-guidelines for musculoskeletal and inflammatory conditions are negligible (< 1.5 degrees C) in light, medium, and dark skin. However, higher LLLT doses delivered with a strong 3B laser (200mW) are capable of increasing skin temperature significantly and these photothermal effects may exceed the thermal pain threshold for humans with dark skin color.

The Importance of Protein-Protein Interactions on the pH-Induced Conformational Changes of Bovine Serum Albumin: A Small-Angle X-Ray Scattering Study

The combined effects of concentration and pH on the conformational states of bovine serum albumin (BSA) are investigated by small-angle x-ray scattering. Serum albumins, at physiological conditions, are found at concentrations of similar to 35-45 mg/mL (42 mg/mL in the case of humans). In this work, BSA at three different concentrations (10, 25, and 50 mg/mL) and pH values (2.0-9.0) have been studied. Data were analyzed by means of the Global Fitting procedure, with the protein form factor calculated from human serum albumin (HSA) crystallographic structure and the interference function described, considering repulsive and attractive interaction potentials within a random phase approximation. Small-angle x-ray scattering data show that BSA maintains its native state from pH 4.0 up to 9.0 at all investigated concentrations. A pH-dependence of the absolute net protein charge is shown and the charge number per BSA is quantified to 10(2), 8(l), 13(2), 20(2), and 26(2) for pH values 4.0, 5.4, 7.0, 8.0, and 9.0, respectively. The attractive potential diminishes as BSA concentration increases. The coexistence of monomers and dimers is observed at 50 mg/mL and pH 5.4, near the BSA isoelectric point. Samples at pH 2.0 show a different behavior, because BSA overall shape changes as a function of concentration. At 10 mg/mL, BSA is partially unfolded and a strong repulsive protein-protein interaction occurs due to the high amount of exposed charge. At 25 and 50 mg/mL, BSA undergoes some refolding, which likely results in a molten-globule state. This work concludes by confirming that the protein concentration plays an important role on the pH-unfolded BSA state, due to a delicate compromise between interaction forces and crowding effects.

Recombinant human prolactin promotes human beta cell survival via inhibition of extrinsic and intrinsic apoptosis pathways

Transplantation of pancreatic islets constitutes a promising alternative treatment for type 1 diabetes. However, it is limited by the shortage of organ donors. Previous results from our laboratory have demonstrated beneficial effects of recombinant human prolactin (rhPRL) treatment on beta cell cultures. We therefore investigated the role of rhPRL action in human beta cell survival, focusing on the molecular mechanisms involved in this process. Human pancreatic islets were isolated using an automated method. Islet cultures were pre-treated in the absence or presence of rhPRL and then subjected to serum starvation or cytokine treatment. Beta cells were labelled with Newport green and apoptosis was evaluated using flow cytometry analysis. Levels of BCL2 gene family members were studied by quantitative RT-PCR and western blot. Caspase-8, -9 and -3 activity, as well as nitric oxide production, were evaluated by fluorimetric assays. The proportion of apoptotic beta cells was significantly lowered in the presence of rhPRL under both cell death-induced conditions. We also demonstrated that cytoprotection may involve an increase of BCL2/BAX ratio, as well as inhibition of caspase-8, -9 and -3. Our study provides relevant evidence for a protective effect of lactogens on human beta cell apoptosis. The results also suggest that the improvement of cell survival may involve, at least in part, inhibition of cell death pathways controlled by the BCL2 gene family members. These findings are highly relevant for improvement of the islet isolation procedure and for clinical islet transplantation.

Human multipotent adipose-derived stem cells restore dystrophin expression of Duchenne skeletal-muscle cells in vitro

Background information. DMD (Duchenne muscular dystrophy) is a devastating X-linked disorder characterized by progressive muscle degeneration and weakness. The use of cell therapy for the repair of defective muscle is being pursued as a possible treatment for DMD. Mesenchymal stem cells have the potential to differentiate and display a myogenic phenotype in vitro. Since liposuctioned human fat is available in large quantities, it may be an ideal source of stem cells for therapeutic applications. ASCs (adipose-derived stem cells) are able to restore dystrophin expression in the muscles of mdx (X-linked muscular dystrophy) mice. However, the outcome when these cells interact with human dystrophic muscle is still unknown. Results. We show here that ASCs participate in myotube formation when cultured together with differentiating human DMD myoblasts, resulting in the restoration of dystrophin expression. Similarly, dystrophin was induced when ASCs were co-cultivated with DMD myotubes. Experiments with GFP (green fluorescent protein)-positive ASCs and DAPI (4,6-diamidino-2-phenylindole)-stained DMD myoblasts indicated that ASCs participate in human myogenesis through cellular fusion. Conclusions. These results show that ASCs have the potential to interact with dystrophic muscle cells, restoring dystrophin expression of DMD cells in vitro. The possibility of using adipose tissue as a source of stem cell therapies for muscular diseases is extremely exciting.

Fatty acids decrease catalase activity in human leukaemia cell lines

Fatty acid (FA) may disturb the redox state of the cells not only by an increase in reactive oxygen species (ROS) generation but also due to a reduction in antioxidant enzyme activities. The effect of various FAs (palmitic, stearic, oleic, linoleic, gamma-linolenic and eicosapentaenoic acids (EPAs)) on Jurkat and Raji cells, (human T and B leukaemic cell lines was investigated). The following measurements were carried out: FA composition of the cells, cell proliferation and activities of catalase, glutathione peroxidase (GPx) and superoxide dismutase (SOD). The protective effect of alpha-tocopherol on cell death was also investigated. Each cell line presented a specific FA composition. All the tested ENS reduced catalase activity. The toxic effect of FA was abolished by the pre-incubation with physiological concentrations of alpha-tocopherol. The findings support the proposition that the increase in oxidative stress induced by FA partially occurs due to a reduction in catalase activity. In spite of the decrease in the enzyme activity, catalase protein and mRNA levels were not changed, suggesting a post-translational regulation. Copyright (C) 2007 John Wiley & Sons, Ltd.

Heat and exercise acclimation increases intracellular levels of Hsp72 and inhibits exercise-induced increase in intracellular and plasma Hsp72 in humans

In order to verify the effects of heat and exercise acclimation (HA) on resting and exercise-induced expression of plasma and leukocyte heat shock protein 72 (Hsp72) in humans, nine healthy young male volunteers (25.0 +/- 0.7 years; 80.5 +/- 2.0 kg; 180 +/- 2 cm, mean +/- SE) exercised for 60 min in a hot, dry environment (40 +/- 0A degrees C and 45 A +/- 0% relative humidity) for 11 days. The protocol consisted of running on a treadmill using a controlled hyperthermia technique in which the work rate was adjusted to elevate the rectal temperature by 1A degrees C in 30 min and maintain it elevated for another 30 min. Before and after the HA, the volunteers performed a heat stress test (HST) at 50% of their individual maximal power output for 90 min in the same environment. Blood was drawn before (REST), immediately after (POST) and 1 h after (1 h POST) HST, and plasma and leukocytes were separated and stored. Subjects showed expected adaptations to HA: reduced exercise rectal and mean skin temperatures and heart rate, and augmented sweat rate and exercise tolerance. In HST1, plasma Hsp72 increased from REST to POST and then returned to resting values 1 h POST (REST: 1.11 A +/- 0.07, POST: 1.48 A +/- 0.10, 1 h POST: 1.22 A +/- 0.11 ng mL(-1); p < 0.05). In HST2, there was no change in plasma Hsp72 (REST: 0.94 A +/- 0.08, POST: 1.20 A +/- 0.15, 1 h POST: 1.17 A +/- 0.16 ng mL(-1); p > 0.05). HA increased resting levels of intracellular Hsp72 (HST1: 1 A +/- 0.02 and HST2: 4.2 A +/- 1.2 density units, p < 0.05). Exercise-induced increased intracellular Hsp72 expression was observed on HST1 (HST1: REST, 1 A +/- 0.02 vs. POST, 2.9 A +/- 0.9 density units, mean +/- SE, p < 0.05) but was inhibited on HST2 (HST2: REST, 4.2 +/- 1.2 vs. POST, 4.4 +/- 1.1 density units, p > 0.05). Regression analysis showed that the lower the pre-exercise expression of intracellular Hsp72, the higher the exercise-induced increase (R = -0.85, p < 0.05). In conclusion, HA increased resting leukocyte Hsp72 levels and inhibited exercise-induced expression. This intracellular adaptation probably induces thermotolerance. In addition, the non-increase in plasma Hsp72 after HA may be related to lower stress at the cellular level in the acclimated individuals.

High frequency tendon reflexes in the human soleus muscle

Tendon reflexes have been often used in studies of the human nervous system in health and disease. They have been investigated either in response to single tendon taps or to long duration vibrations. Tendon reflexes are described here in response to a high frequency vibration burst (3 cycles of a 100 Hz sine wave) applied to the Achilles tendon of standing subjects, either in quiet stance or during a forward leaning posture. The electromyogram from the soleus muscle usually showed three components separated by 10 ms which were interpreted as being three reflexes, each reflex induced by each of the three cycles in a burst. This result indicates that soleus tendon reflexes can respond in fast succession in a phasic manner when a brief high frequency vibration is applied to the Achilles tendon. This occurs in spite of possible depression of the la to motoneuron synapses and the long after hyperpolarization of the motoneurons. An interpretation of the results is that motoneurons from different subsets of the motoneuron pool respond to different cycles of the sinusoidal vibratory burst. (c) 2008 Elsevier Ireland Ltd. All rights reserved.