Human Fallopian Tube Mesenchymal Stromal Cells Enhance Bone Regeneration in a Xenotransplanted Model

We have recently reported that human fallopian tubes, which are discarded during surgical procedures of women submitted to sterilization or hysterectomies, are a rich source of human fallopian tube mesenchymal stromal cells (htMSCs). It has been previously shown that human mesenchymal stromal cells may be useful in enhancing the speed of bone regeneration. This prompted us to investigate whether htMSCs might be useful for the treatment of osteoporosis or other bone diseases, since they present a pronounced capacity for osteogenic differentiation in vitro. Based on this prior knowledge, our aim was to evaluate, in vivo, the osteogenic capacity of htMSCs to regenerate bone through an already described xenotransplantation model: nonimmunosuppressed (NIS) rats with cranial defects. htMSCs were obtained from five 30-50 years old healthy women and characterized by flow cytometry and for their multipotenciality in vitro capacity (osteogenic, chondrogenic and adipogenic differentiations). Two symmetric full-thickness cranial defects on each parietal region of seven NIS rats were performed. The left side (LS) of six animals was covered with CellCeram (Scaffdex)-a bioabsorbable ceramic composite scaffold that contains 60% hydroxyapatite and 40% beta-tricalciumphosphate-only, and the right side (RS) with the CellCeram and htMSCs (10(6) cells/scaffold). The animals were euthanized at 30, 60 and 90 days postoperatively and cranial tissue samples were taken for histological analysis. After 90 days we observed neobone formation in both sides. However, in animals euthanized 30 and 60 days after the procedure, a mature bone was observed only on the side with htMSCs. PCR and immunofluorescence analysis confirmed the presence of human DNA and thus that human cells were not rejected, which further supports the imunomodulatory property of htMSCs. In conclusion, htMSCs can be used successfully to enhance bone regeneration in vivo, opening a new field for future treatments of osteoporosis and bone reconstruction.

Local Injections of Adipose-Derived Mesenchymal Stem Cells Modulate Inflammation and Increase Angiogenesis Ameliorating the Dystrophic Phenotype in Dystrophin-Deficient Skeletal Muscle

The effects of adipose-derived mesenchymal stem cells (ADMSC) transplantation on degeneration, regeneration and skeletal muscle function were investigated in dystrophin-deficient mice (24-week-old). ADMSC transplantation improved muscle strength and, resistance to fatigue. An increase in fiber cross-sectional area and in the number of fibers with centralized nuclei and augment of myogenin content were observed. In ADMSC-treated muscles a decrease in muscle content of TNF-alpha, IL-6 and oxidative stress measured by Amplex(A (R)) reagent were observed. The level of TGF-beta 1 was lowered whereas that of VEGF, IL-10 and IL-4 were increased by ADMSC treatment. An increase in markers of macrophage M1 (CD11 and F4-80) and a decrease in T lymphocyte marker (CD3) and arginase-1 were also observed in ADMSCs-treated dystrophic muscle. No change was observed in iNOS expression. Increased phosphorylation of Akt, p70S6k and 4E-BP1 was found in dystrophic muscles treated with ADMSC. These results suggest that ADMSC transplantation modulates inflammation and improves muscle tissue regeneration, ameliorating the dystrophic phenotype in dystrophin-deficient mice.

Monoclonal B-cell lymphocytosis (MBL), CD4(+)/CD8(weak) T-cell large granular lymphocytic leukemia (T-LGL leukemia) and monoclonal gammopathy of unknown significance (MGUS): molecular and flow cytometry characterization of three concomitant hematological disorders

The diagnosis of T-cell large granular lymphocytic leukemia in association with other B-cell disorders is uncommon but not unknown. However, the concomitant presence of three hematological diseases is extraordinarily rare. We report an 88-year-old male patient with three simultaneous clonal disorders, that is, CD4+/CD8(weak) T-cell large granular lymphocytic leukemia, monoclonal gammopathy of unknown significance and monoclonal B-cell lymphocytosis. The patient has only minimal complaints and has no anemia, neutropenia or thrombocytopenia. Lymphadenopathy and hepatosplenomegaly were not present. The three disorders were characterized by flow cytometry analysis, and the clonality of the T-cell large granular lymphocytic leukemia was confirmed by polymerase chain reaction. Interestingly, the patient has different B-cell clones, given that plasma cells of monoclonal gammopathy of unknown significance exhibited a kappa light-chain restriction population and, on the other hand, B-lymphocytes of monoclonal B-cell lymphocytosis exhibited a lambda light-chain restriction population. This finding does not support the antigen-driven hypothesis for the development of multi-compartment diseases, but suggests that T-cell large granular lymphocytic expansion might represent a direct antitumor immunological response to both B-cell and plasma-cell aberrant populations, as part of the immune surveillance against malignant neoplasms.

Effect of Substrate Concentration on Dark Fermentation Hydrogen Production Using an Anaerobic Fluidized Bed Reactor

The effect of substrate (glucose) concentration on the stability and yield of a continuous fermentative process that produces hydrogen was studied. Four anaerobic fluidized bed reactors (AFBRs) were operated with a hydraulic retention time (HRT) from 1 to 8 h and an influent glucose concentration from 2 to 25 gL(-1). The reactors were inoculated with thermally pre-treated anaerobic sludge and operated at a temperature of 30 degrees C with an influent pH around 5.5 and an effluent pH of about 3.5. The AFBRs with a HRT of 2 h and a feed strength of 2, 4, and 10 gL(-1) showed satisfactory H-2 production performance, but the reactor fed with 25 gL(-1) of glucose did not. The highest hydrogen yield value was obtained in the reactor with a glucose concentration of 2 gL(-1) when it was operated at a HRT of 2 h. The maximum hydrogen production rate value was achieved in the reactor with a HRT of 1 h and a feed strength of 10 gL(-1). The AFBRs operated with glucose concentrations of 2 and 4 gL(-1) produced greater amounts of acetic and butyric acids, while AFBRs with higher glucose concentrations produced a greater amount of solvents.

Morphological Homogeneity of Neurons: Searching for Outlier Neuronal Cells

We report a morphology-based approach for the automatic identification of outlier neurons, as well as its application to the NeuroMorpho.org database, with more than 5,000 neurons. Each neuron in a given analysis is represented by a feature vector composed of 20 measurements, which are then projected into a two-dimensional space by applying principal component analysis. Bivariate kernel density estimation is then used to obtain the probability distribution for the group of cells, so that the cells with highest probabilities are understood as archetypes while those with the smallest probabilities are classified as outliers. The potential of the methodology is illustrated in several cases involving uniform cell types as well as cell types for specific animal species. The results provide insights regarding the distribution of cells, yielding single and multi-variate clusters, and they suggest that outlier cells tend to be more planar and tortuous. The proposed methodology can be used in several situations involving one or more categories of cells, as well as for detection of new categories and possible artifacts.

Response of Saccharomyces cerevisiae to Cadmium and Nickel Stress: The Use of the Sugar Cane Vinasse as a Potential Mitigator

Most of the metals released from industrial activity, among them are cadmium (Cd) and nickel (Ni), inhibit the productivity of cultures and affect microbial metabolism. In this context, the aim of this work was to investigate the capacity of sugar cane vinasse to mitigate the adverse effects of Cd and Ni on cell growth, viability, budding rate and trehalose content of Saccharomyces cerevisiae, likely because of adsorption and chelating action. For this purpose, the yeast was grown batch-wise in YED medium supplemented with selected amounts of vinasse and Cd or Ni. The negative effects of Cd and Ni on S. cerevisiae growth and the mitigating one of sugar cane vinasse were quantified by an exponential model. Without vinasse, the addition of increasing levels of Cd and Ni reduced the specific growth rate, whereas in its presence no reduction was observed. Consistently with the well-proved toxicity of both metals, cell viability and budding rate progressively decreased with increasing their concentration, but in the presence of vinasse the situation was remarkably improved. The trehalose content of S. cerevisiae cells followed the same qualitative behavior as cell viability, even though the negative effect of both metals on this parameter was stronger. These results demonstrate the ability of sugar cane vinasse to mitigate the toxic effects of Cd and Ni.

Xylanase and beta-Xylosidase Production by Aspergillus ochraceus: New Perspectives for the Application of Wheat Straw Autohydrolysis Liquor

The xylanase biosynthesis is induced by its substrate-xylan. The high xylan content in some wastes such as wheat residues (wheat bran and wheat straw) makes them accessible and cheap sources of inducers to be mainly applied in great volumes of fermentation, such as those of industrial bioreactors. Thus, in this work, the main proposal was incorporated in the nutrient medium wheat straw particles decomposed to soluble compounds (liquor) through treatment of lignocellulosic materials in autohydrolysis process, as a strategy to increase and undervalue xylanase production by Aspergillus ochraceus. The wheat straw autohydrolysis liquor produced in several conditions was used as a sole carbon source or with wheat bran. The best conditions for xylanase and beta-xylosidase production were observed when A. ochraceus was cultivated with 1% wheat bran added of 10% wheat straw liquor (produced after 15 min of hydrothermal treatment) as carbon source. This substrate was more favorable when compared with xylan, wheat bran, and wheat straw autohydrolysis liquor used separately. The application of this substrate mixture in a stirred tank bioreactor indicated the possibility of scaling up the process to commercial production.

A new dic(7;12)(p12.21;p12.2) and i(12)(q10) during the lymphoid blast crisis of patient with Ph plus chronic myeloid leukemia

Chronic myelogenous leukemia (CML) is a common myeloproliferative disease that is characterized by the clonal expansion of marrow stem cells, and is associated with the Philadelphia chromosome. As the disease progresses, additional chromosome abnormalities may arise. The prognostic impact of secondary chromosomal abnormalities in CML is complex, heterogeneous, and sometimes related to previous treatment. Here, we describe a CML patient in lymphoid blast crisis associated with a new chromosomal abnormality identified, dic(7;12)(p12.21;p12.2) and i(12)(q10) using classical cytogenetics and spectral karyotype analysis. To the best of our knowledge, this is the first report of t(7;12)(p11.1;q11.1) and i(12)(q10) in a CML patient with lymphoid evolution.

Acute promyelocytic leukemia associated with the PLZF-RARA fusion gene: two additional cases with clinical and laboratorial peculiar presentations

Acute promyelocytic leukemia (APL) is characterized by the presence of the t(15;17) and PML-RARa rearrangement, with good response to treatment with retinoids. However, few cases of variant APL involving alternative chromosomal aberrations have been reported, including t(11;17)(q23;q21) (Wells et al. in Nat Genet 17:109-113, 1; Arnould et al. in Hum Mol Genet 8:1741-1749, 2) t(5;17)(q35;q12-21), t(11;17)(q13;q21) (Grimwade et al in Blood 96:1297-1308, 3) and der(17) (Rego et al. in Blood (ASH Annual Meeting Abstracts)114:Abstract 6, 4), whereby RARa is fused to the PLZF, NPM, NuMA, and STAT5b genes, respectively, have been described. These cases are characterized by distinct morphology, clinical presentation, and in respect to PLZF, a lack of differentiation response to retinoids leading to the need of different approaches concerning diagnostic methods and therapeutics. This paper describes two cases of APL associated with the PLZF-RARA fusion gene enrolled in the IC-APL trial that is a non-randomized, multicenter study conducted in Brazil, Mexico, Chile and Uruguay with the aim to improve the treatment outcome of APL patients in developing countries. These cases, although rare, offer a challenge to its early recognition and proper conduction.

Preservation of Alpha-3 Neuronal Nicotinic Acetylcholine Receptor Expression in Sympathetic Ganglia After Brain Death

The goal of this study was to evaluate if the immunohistochemical expression of alpha-3 neuronal nicotinic acetylcholine receptor subunit in sympathetic ganglia remains stable after brain death, determining the possible use of sympathetic thoracic ganglia from subjects after brain death as study group. The third left sympathetic ganglion was resected from patients divided in two groups: BD-organ donors after brain death and CON-patients submitted to sympathectomy for hyperhidrosis (control group). Immunohistochemical staining for alpha-3 neuronal nicotinic acetylcholine receptor subunit was performed; strong and weak expression areas were quantified in both groups. The BD group showed strong alpha-3 neuronal nicotinic acetylcholine receptor expression in 6.55% of the total area, whereas the CON group showed strong expression in 5.91% (p = 0.78). Weak expression was found in 6.47% of brain-dead subjects and in 7.23% of control subjects (p = 0.31). Brain death did not affect the results of the immunohistochemical analysis of sympathetic ganglia, and its use as study group is feasible.

A Two-Stage Aerobic/Anaerobic Denitrifying Horizontal Bioreactor Designed for Treating Ammonium and H2S Simultaneously

A two-stage bioreactor was operated for a period of 140 days in order to develop a post-treatment process based on anaerobic bioxidation of sulfite. This process was designed for simultaneously treating the effluent and biogas of a full-scale UASB reactor, containing significant concentrations of NH4 and H2S, respectively. The system comprised of two horizontal-flow bed-packed reactors operated with different oxygen concentrations. Ammonium present in the effluent was transformed into nitrates in the first aerobic stage. The second anaerobic stage combined the treatment of nitrates in the liquor with the hydrogen sulfide present in the UASB-reactor biogas. Nitrates were consumed with a significant production of sulfate, resulting in a nitrate removal rate of 0.43 kg N m(3) day(-1) and a parts per thousand yen92 % efficiency. Such a removal rate is comparable to those achieved by heterotrophic denitrifying systems. Polymeric forms of sulfur were not detected (elementary sulfur); sulfate was the main product of the sulfide-based denitrifying process. S-sulfate was produced at a rate of about 0.35 kg m(3) day(-1). Sulfur inputs as S-H2S were estimated at about 0.75 kg m(3) day(-1) and Chemical Oxygen Demand (COD) removal rates did not vary significantly during the process. DGGE profiling and 16S rRNA identified Halothiobacillus-like species as the key microorganism supporting this process; such a strain has not yet been previously associated with such bioengineered systems.

Hydrogen and Methane Production, Energy Recovery, and Organic Matter Removal from Effluents in a Two-Stage Fermentative Process

This study evaluates the potential for using different effluents for simultaneous H-2 and CH4 production in a two-stage batch fermentation process with mixed microflora. An appreciable amount of H-2 was produced from parboiled rice wastewater (23.9 mL g(-1) chemical oxygen demand [COD]) and vinasse (20.8 mL g(-1) COD), while other effluents supported CH4 generation. The amount of CH4 produced was minimum for sewage (46.3 mL g(-1) COD), followed by parboiled rice wastewater (115.5 mL g(-1) COD) and glycerol (180.1 mL g(-1) COD). The maximum amount of CH4 was observed for vinasse (255.4 mL g(-1) COD). The total energy recovery from vinasse (10.4 kJ g(-1) COD) corresponded to the maximum COD reduction (74.7 %), followed by glycerol (70.38 %, 7.20 kJ g(-1) COD), parboiled rice wastewater (63.91 %, 4.92 kJ g(-1) COD), and sewage (51.11 %, 1.85 kJ g(-1) COD). The relatively high performance of vinasse in such comparisons could be attributed to the elevated concentrations of macronutrients contained in raw vinasse. The observations are based on kinetic parameters of H-2 and CH4 production and global energy recovery of the process. These observations collectively suggest that organic-rich effluents can be deployed for energy recovery with sequential generation of H-2 and CH4.

Biotechnological Utilization of Biodiesel-Derived Glycerol for the Production of Ribonucleotides and Microbial Biomass

Ten yeast strains were evaluated concerning their capabilities to assimilate biodiesel-derived glycerol in batch cultivation. The influence of glycerol concentration, temperature, pH and yeast extract concentration on biomass production was studied for the yeast selected. Further, the effect of agitation on glycerol utilization by the yeast Hansenula anomala was also studied. The yeast H. anomala CCT 2648 showed the highest biomass yield (0.30 g g(-1)) and productivity (0.19 g L-1 h(-1)). Citric acid, succinic acid, acetic acid and ethanol were found as the main metabolites produced. The increase of yeast extract concentration from 1 to 3 g L-1 resulted in high biomass production. The highest biomass concentration (21 g L-1), yield (0.45 g g(-1)) and productivity (0.31 g L-1 h(-1)), as well as ribonucleotide production (13.13 mg g(-1)), were observed at 700 rpm and 0.5 vvm. These results demonstrated that glycerol from biodiesel production process showed to be a feasible substrate for producing biomass and ribonucleotides by yeast species.

CD44(+)/CD24(-) cells and lymph node metastasis in stage I and II invasive ductal carcinoma of the breast

The presence of tumor-initiating cells (CD44(+)/CD24(-)) in solid tumors has been reported as a possible cause of cancer metastasis and treatment failure. Nevertheless, little is know about the presence of CD44(+)/CD24(-) cells within the primary tumor and metastasis. The proportion of CD44(+)/CD24(-) cells was analyzed in 40 samples and in 10 lymph node metastases using flow cytometry phenotyping. Anti-human CD326 (EpCam; FITC), antihuman CD227 (MUC-1; FITC), anti-human CD44 (APC), and anti-human CD24 (PE), anti-ABCG2 (PE), and anti-CXCR4 (PeCy7) were used for phenotype analysis. The mean patient age was 60.5 years (range, 33-87 years); mean primary tumor size (pT) was 1.8 cm (0.5-3.5 cm). The Wilcoxon or Kruskal-Wallis test was used for univariate analyses. Logistic regression was used for multivariate analysis. The median percentage of CD44(+)/CD24(-) cells within primary invasive ductal carcinomas (IDC) was 2.7% (range, 0.2-71.2). In lymph node metastases, we observed a mean of 6.1% (range, 0.07-53.7). The percentage of CD44(+)/CD24(-) cells in IDCs was not associated with age, pT, tumor grade and HER2. We observed a significantly enrichment of CD44(+)/CD24(-) and ABCG2(+) cells in ESA(+) cell population in patients with positive lymph nodes (P = 0.02 and P = 0.04, respectively). Our data suggest that metastatic dissemination is associated with an increase in tumorinitiating cells in stage I and II breast cancer.