Callus induction and micropropagation improved by colchicine and phytoregulators in Kappaphycus alvarezii (Rhodophyta, Solieriaceae)

Tissue culture techniques were applied for micropropagation of the red alga Kappaphycus alvarezii in order to select the best strain and experimental system for in vitro culture. Five strains were tested: brown (BR), green (GR) and red (RD) tetrasporophytes, brown female gametophyte (BFG), and a strain originating from tetraspore germination (""Edison de Paula"", EP). The effects of three culture media were tested on callus formation, regeneration from explants and from callus in the three tetrasporophytic and EP strains: seawater enriched with half-strength of von Stosch`s (VS 50) and Guillard & Ryther`s (F/2 50) solutions, plus synthetic ASP 12-NTA medium, with or without gelling agent. Explants of the EP strain were treated with glycerol and the phytoregulators indole-3-acetic acid (IAA); 2,4-diclorophenoxyacetic acid (2,4-D); and benzylaminopurine (BA), alone or in combination. The effects of colchicine (0.01%) during 24, 48, 72 hours and 14 days were analyzed in the BFG and EP strains. The EP strain showed the highest percentage of explants forming callus and regeneration from explants in VS 50, indicating its high potential for micropropagation in comparison to the other strains. Regeneration from callus was very rare. Treatments with glycerol and IAA:BA (5:1 mg L(-1)) stimulated the regeneration from explants. Significant differences were observed in the percentages of regeneration of EP strain explants treated with colchicine for 14 days. Our results indicate that IAA and BA stimulated the regeneration process, and that colchicine produced explants with high potential for regeneration, being useful for improving the micropropagation of K. alvarezii.

In vitro morphogenesis and cell suspension culture establishment in Piper solmsianum C. DC. (Piperaceae)

In vitro morphogenesis and cell suspension culture establishment in Piper solmsianum C. DC. (Piperaceae)). Piper solmsianum is a shrub from Southeast Brazil in which many biologically active compounds were identified. The aim of this work was to establish a cell suspension culture system for this species. With this in mind, petiole and leaf explants obtained from in vitro plantlets were cultured in the presence of different plant growth regulator combinations (IAA, NAA, 2,4-D and BA). Root and indirect shoot adventitious formation, detected by histological analysis, was observed. Besides the different combinations of plant growth regulators, light regime and the supplement of activated charcoal (1.5 mg.l(-1)) were tested for callus induction and growth. Cultures maintained in light, on a 0.2 mg.l(-1) 2,4-D and 2 mg.l(-1) BA supplemented medium, and in the absence of activated charcoal, showed the highest calli fresh matter increment. From a callus culture, cell suspension cultures were established and their growth and metabolite accumulation studied. The achieved results may be useful for further characterization of the activated secondary metabolites pathways in in vitro systems of P. solmsianum.

Histopathological evaluation of different methods of experimental induction of periapical periodontitis

This study evaluated histopathologically different methods of experimental induction of periapical periodontitis. The radiographic and microbiological evaluations have been performed in a previous investigation. Fifty-seven root canals from dogs' teeth were assigned to 4 groups. In GI (n=14) and GII (n=14), the root canals were exposed to oral environment for 180 days; in GIII (n=14) and GIV (n=15) the root canals were exposed for 7 days and then the access cavities were restored and remained sealed for 53 days. The root apices of GI and GIII were perforated, whilst those of GII and GIV remained intact. After induction of periapical periodontitis, the dogs were euthanized. Serial sections were obtained and stained with hematoxylin and eosin. Data of the histopathological evaluation were submitted to Kruskal-Wallis and Dunn's tests at 5% significance level. The inflammatory periapical reaction and resorption of mineralized tissues were less intense in GII than in the other groups (p<0.05). There was no histopathological difference among the experimentally induced periapical lesions in the teeth with coronal sealing. On the other hand, when coronal sealing was not performed, greater intensity of induced periapical periodontitis was observed in the teeth with apical perforation.

Microbial distribution in the root canal system after periapical lesion induction using different methods

The aim of this study was to evaluate the microbial distribution in the root canal system after periapical lesion induction in dogs' teeth using different methods. Fifty-two root canals were assigned to 4 groups (n=13). Groups I and II: root canals were exposed to the oral cavity for 180 days; groups III and IV: root canals were exposed for 7 days and then the coronal openings were sealed for 53 days. The root apices of groups I and III were perforated, while those of groups II and IV remained intact. After the experimental periods, the animals were euthanized and the anatomic pieces containing the roots were processed and stained with the Brown & Brenn method to assess the presence and distribution of microorganisms. The incidence of microorganisms at different sites of the roots and periapical lesions was analyzed statistically by the chi-square test at 5% significance level. All groups presented microorganisms in the entire root canal system. A larger number of microorganisms was observed on the root canal walls, apical delta and dentinal tubules (p<0.05), followed by cementum and cemental resorption areas. In spite of the different periods of exposure to the oral environment, the methods used for induction of periapical periodontitis yielded similar distribution of microorganisms in the root canal system.

Efeitos do ágar no crescimento de explantes e na formação de calos em morfos pigmentares de Gracilaria domingensis (Kützing) Sonder ex Dickie (Gracilariales, Rhodophyta)

Os efeitos da concentração de ágar no crescimento de explantes e na formação de calos foram avaliados em culturas axênicas de gametófitos femininos de morfos de coloração verde e vermelha de Gracilaria domingensis (Kützing) Sonder ex Dickie. Culturas unialgáceas foram mantidas em água do mar esterilizada (30-32 ups) enriquecida com 25% da solução de von Stosch (VSES 25%), 22 ± 2 °C, fotoperíodo de 14 h, irradiância de 50-80 µmol de fótons m-2 s-1. Para a obtenção de explantes axênicos, segmentos apicais e intercalares dos dois morfos foram cultivados por 48 h em meio VSES 25% com adição de uma solução antibiótica e antifúngica, e submetidos a uma lavagem com uma solução de água do mar esterilizada com 0,5% de hipoclorito de sódio e 200 µL L-1 de detergente por 20 segundos. Para avaliar os efeitos da concentração de ágar, os segmentos axênicos foram inoculados em meio ASP 12-NTA com concentrações distintas de ágar que variaram de zero a 1%. A adição de ágar no meio inibiu o crescimento dos segmentos apicais de ambos os morfos, bem como o crescimento de segmentos intercalares do morfo verde. Observou-se uma tendência geral no crescimento dos explantes, onde a taxa de crescimento foi inversamente proporcional à concentração de ágar. A adição de ágar no meio induziu a formação de três tipos de calo, denominados conforme a região do explante onde se originaram: calo apical, calo basal e calo intermediário. As concentrações de 0,5% e 0,7% de ágar foram as concentrações ótimas para indução de calos basais e calos intermediários no morfo verde, respectivamente. A presença de ágar foi essencial para a formação de calos intermediários e apicais. Os resultados indicam que o ágar apresenta um papel na regulação dos processos morfogenéticos em morfos pigmentares de G. domingensis.

Cognitive control associated with irritability induction: an autobiographical recall fMRI study

OBJECTIVE: Despite the relevance of irritability emotions to the treatment, prognosis and classification of psychiatric disorders, the neurobiological basis of this emotional state has been rarely investigated to date. We assessed the brain circuitry underlying personal script-driven irritability in healthy subjects (n = 11) using functional magnetic resonance imaging. METHOD: Blood oxygen level-dependent signal changes were recorded during auditory presentation of personal scripts of irritability in contrast to scripts of happiness or neutral emotional content. Self-rated emotional measurements and skin conductance recordings were also obtained. Images were acquired using a 1,5T magnetic resonance scanner. Brain activation maps were constructed from individual images, and between-condition differences in the mean power of experimental response were identified by using cluster-wise nonparametric tests. RESULTS: Compared to neutral scripts, increased blood oxygen level-dependent signal during irritability scripts was detected in the left subgenual anterior cingulate cortex, and in the left medial, anterolateral and posterolateral dorsal prefrontal cortex (cluster-wise p-value < 0.05). While the involvement of the subgenual cingulate and dorsal anterolateral prefrontal cortices was unique to the irritability state, increased blood oxygen level-dependent signal in dorsomedial and dorsal posterolateral prefrontal regions were also present during happiness induction. CONCLUSION: Irritability induction is associated with functional changes in a limited set of brain regions previously implicated in the mediation of emotional states. Changes in prefrontal and cingulate areas may be related to effortful cognitive control aspects that gain salience during the emergence of irritability.

Induction of Zenk protein expression within the nucleus taeniae of the amygdala of pigeons following tone and shock stimulation

In this study, we evaluated the expression of the Zenk protein within the nucleus taeniae of the pigeon’s amygdala (TnA) after training in a classical aversive conditioning, in order to improve our understanding of its functional role in birds. Thirty-two 18-month-old adult male pigeons (Columba livia), weighing on average 350 g, were trained under different conditions: with tone-shock associations (experimental group; EG); with shock-alone presentations (shock group; SG); with tone-alone presentations (tone group; TG); with exposure to the training chamber without stimulation (context group; CG), and with daily handling (naive group; NG). The number of immunoreactive nuclei was counted in the whole TnA region and is reported as density of Zenk-positive nuclei. This density of Zenk-positive cells in the TnA was significantly greater for the EG, SG and TG than for the CG and NG (P < 0.05). The data indicate an expression of Zenk in the TnA that was driven by experience, supporting the role of this brain area as a critical element for neural processing of aversive stimuli as well as meaningful novel stimuli.

Induction of a gloverin-like antimicrobial polypeptide in the sugarcane borer Diatraea saccharalis challenged by septic injury

Diatraea saccharalis (Fabricius, 1794) (Lepidoptera: Crambidae) is an important pest for Brazilian sugarcane. In the present study, we detected two distinct spots in hemolymph from septic injured larvae (HDs1 and HDs2), which are separated by 2DE gel electrophoresis. Both spots were subjected to in-gel tryptic digestion and MALDI-TOF/TOF analysis, which revealed the sequence VFGTLGSDDSGLFGK present in both HDs1 and HDs2. This sequence had homology and 80% identity with specific Lepidoptera antimicrobial peptides called gloverins. Analyses using the ImageMaster 2D software showed pI 8.94 of the HDs1 spot, which is similar to that described to Hyalophora gloveri gloverin (pI 8.5). Moreover, the 14-kDa molecular mass of the spot HDs1 is compatible to that of gloverins isolated from the hemolymph of Trichoplusia ni, Helicoverpa armigera and H. gloveri. Antimicrobial assays with partially purified fractions containing the HDs1 and HDs2 polypeptides demonstrated activity against Escherichia coli. This is the first report of antimicrobial polypeptides in D. saccharalis, and the identification of these peptides may help in the generation of new strategies to control this pest.

Crowbar System in Doubly Fed Induction Wind Generators

In the last 15 years, the use of doubly fed induction machines in modern variable-speed wind turbines has increased rapidly. This development has been driven by the cost reduction as well as the low-loss generation of Insulated Gate Bipolar Transistors (IGBT). According to new grid code requirements, wind turbines must remain connected to the grid during grid disturbances. Moreover, they must also contribute to voltage support during and after grid faults. The crowbar system is essential to avoid the disconnection of the doubly fed induction wind generators from the network during faults. The insertion of the crowbar in the rotor circuits for a short period of time enables a more efficient terminal voltage control. As a general rule, the activation and the deactivation of the crowbar system is based only on the DC-link voltage level of the back-to-back converters. In this context, the authors discuss the critical rotor speed to analyze the instability of doubly fed induction generators during grid faults.

Resprouting from roots in four Brazilian tree species

Previous studies pointed out that species richness and high density values within the Leguminosae in Brazilian forest fragments affected by fire could be due, at least partially, to the high incidence of root sprouting in this family. However, there are few Studies of the factors that induce root sprouting in woody plants after disturbance. We investigated the bud formation on root cuttings, and considered a man-made disturbance that isolates the root from the shoot apical dominance of three Leguminosae (Bauhinia forficata Link., Centrolobium tomentosum Guill. ex Benth, and Inga laurina (Sw.) Willd) and one Rutaceae (Esenbeckia febrifuga (St. Hit.) Juss. ex Mart.). All these species resprout frequently after fire. We also attempted to induce bud formation on root systems by removing the main trunk, girdling or sectioning the shallow lateral roots from forest tree species Esenbeckia febrifuga and Hymenaea courbaril L. We identified the origin of shoot primordia and their early development by fixing the samples in Karnovsky solution, dehydrating in ethyl alcohol series and embedding in plastic resin. Serial sections were cut on a rotary microtome and stained with toluidine blue O. Permanent slides were mounted in synthetic resin. We observed different modes of bud origin on root cuttings: close to the vascular cambium (C. tomentosum), from the callus (B. forficata and E febrifuga) and from the phloematic parenchyma proliferation (L laurina). Fragments of B. forficala root bark were also capable of forming reparative buds from healing phellogen formed in callus in the bark's inner side. In the attempt of bud induction on root systems, Hymenaea courbaril did not respond to any of the induction tests, probably because of plant age. However, Esenbeckia febrifuga roots formed suckers when the main trunk was removed or their roots were sectioned and isolated from the original plant. We experimentally demonstrated the ability of four tree species to resprout from roots after disturbance. Our results suggest that the release of apical dominance enables root resprouting in the studied species. Rev. Biol. Trop. 57 (3): 789-800. Epub 2009 September 30.

Plasmodium vivax: Induction of CD4(+)CD25(+)FoxP3(+) Regulatory T Cells during Infection Are Directly Associated with Level of Circulating Parasites

Circulation CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) have been associated with the delicate balancing between control of overwhelming acute malaria infection and prevention of immune pathology due to disproportionate inflammatory responses to erythrocytic stage of the parasite. While the role of Tregs has been well-documented in murine models and P. falciparum infection, the phenotype and function of Tregs in P. vivax infection is still poorly characterized. In the current study, we demonstrated that patients with acute P. vivax infection presented a significant augmentation of circulating Tregs producing anti-inflammatory (IL-10 and TGF-beta) as well as pro-inflammatory (IFN-gamma, IL-17) cytokines, which was further positively correlated with parasite burden. Surface expression of GITR molecule and intracellular expression of CTLA-4 were significantly upregulated in Tregs from infected donors, presenting also a positive association between either absolute numbers of CD4(+)CD25(+)FoxP3(+)GITR(+) or CD4(+)CD25(+)FoxP3(+)CTLA-4(+) and parasite load. Finally, we demonstrate a suppressive effect of Treg cells in specific T cell proliferative responses of P. vivax infected subjects after antigen stimulation with Pv-AMA-1. Our findings indicate that malaria vivax infection lead to an increased number of activated Treg cells that are highly associated with parasite load, which probably exert an important contribution to the modulation of immune responses during P. vivax infection.

Induction of Heme Oxygenase-1 Can Halt and Even Reverse Renal Tubule-Interstitial Fibrosis

Background: The tubule-interstitial fibrosis is the hallmark of progressive renal disease and is strongly associated with inflammation of this compartment. Heme-oxygenase-1 (HO-1) is a cytoprotective molecule that has been shown to be beneficial in various models of renal injury. However, the role of HO-1 in reversing an established renal scar has not yet been addressed. Aim: We explored the ability of HO-1 to halt and reverse the establishment of fibrosis in an experimental model of chronic renal disease. Methods: Sprague-Dawley male rats were subjected to unilateral ureteral obstruction (UUO) and divided into two groups: non-treated and Hemin-treated. To study the prevention of fibrosis, animals were pre-treated with Hemin at days -2 and -1 prior to UUO. To investigate whether HO-1 could reverse established fibrosis, Hemin therapy was given at days 6 and 7 post-surgery. After 7 and/or 14 days, animals were sacrificed and blood, urine and kidney tissue samples were collected for analyses. Renal function was determined by assessing the serum creatinine, inulin clearance, proteinuria/creatininuria ratio and extent of albuminuria. Arterial blood pressure was measured and fibrosis was quantified by Picrosirius staining. Gene and protein expression of pro-inflammatory and pro-fibrotic molecules, as well as HO-1 were performed. Results: Pre-treatment with Hemin upregulated HO-1 expression and significantly reduced proteinuria, albuminuria, inflammation and pro-fibrotic protein and gene expressions in animals subjected to UUO. Interestingly, the delayed treatment with Hemin was also able to reduce renal dysfunction and to decrease the expression of pro-inflammatory molecules, all in association with significantly reduced levels of fibrosis-related molecules and collagen deposition. Finally, TGF-beta protein production was significantly lower in Hemin-treated animals. Conclusion: Treatment with Hemin was able both to prevent the progression of fibrosis and to reverse an established renal scar. Modulation of inflammation appears to be the major mechanism behind HO-1 cytoprotection.

Effects of Aflatoxin B(1) and Fumonisin B(1) on the Viability and Induction of Apoptosis in Rat Primary Hepatocytes

The present study evaluated the effect of aflatoxin B(1) (AFB(1)) and fumonisin B(1) (FB(1)) either alone, or in association, on rat primary hepatocyte cultures. Cell viability was assessed by flow cytometry after propidium iodine intercalation. DNA fragmentation and apoptosis were assessed by agarose gel electrophoresis and acridine orange and ethidium bromide staining. At the concentrations of AFB(1) and FB(1) used, the toxins did not decrease cell viability, but did induce apoptosis in a concentration and time-dependent manner.

Induction of mitotic crossing-over in diploid strains of Aspergillus nidulans using low-dose X-rays

As a contribution towards detecting the genetic effects of low doses of genotoxic physical agents, this paper deals with the consequences of low-dose X-rays in the Aspergillus nidulans genome. The irradiation doses studied were those commonly used in dental clinics (1-5 cGy). Even very low doses promoted increased mitotic crossing-over frequencies in diploid strains heterozygous for several genetic markers including the ones involved in DNA repair and recombination mechanisms. Genetic markers of several heterozygous strains were individu`ally analyzed disclosing that some markers were especially sensitive to the treatments. These markers should be chosen as bio-indicators in the homozygotization index assay to better detect the recombinogenic/carcinogenic genomic effects of low-dose X-rays.

Embedded DSP-Based Compact Fuzzy System and Its Application for Induction-Motor V/f Speed Control

This paper presents a compact embedded fuzzy system for three-phase induction-motor scalar speed control. The control strategy consists in keeping constant the voltage-frequency ratio of the induction-motor supply source. A fuzzy-control system is built on a digital signal processor, which uses speed error and speed-error variation to change both the fundamental voltage amplitude and frequency of a sinusoidal pulsewidth modulation inverter. An alternative optimized method for embedded fuzzy-system design is also proposed. The controller performance, in relation to reference and load-torque variations, is evaluated by experimental results. A comparative analysis with conventional proportional-integral controller is also achieved.