Seed abortion in the sexual counterpart of <Emphasis Type="Italic">Brachiaria brizantha</Emphasis> apomicts (Poaceae)

Gametophytic organization, fertilization and reproductive success are described for the fertile diploid Brachiaria brizantha accession BRA-002747 which is being raised for use in Brachiaria breeding programs, as well as to understand and control of apomixis in this genus. The current paper reports on reproductive biology and analysis of seed set in field experiments during three consecutive years. Unsuccessful seed production in this plant is believed to correlate with early inbreeding depression, based on the reproductive features analyzed. Caryopsis development was observed using differential interference contrast microscopy with seed set determined by the number of self- and open-pollinated pistils that fully developed into viable seeds. Developing and mature female and male gametophytes were observed in the context of flower phenology, morphology and anthesis patterns. Pollen viability was determined by acetocarmine staining and by observation of germination in vivo, which was also used to observe pollen tube/pistil interaction. Although normal development was observed in floral structures, anthesis and gametophytes, seed set was low, with 2 and 6% in self- and open-pollination, respectively, producing seed. Variations observed in the female organs, such as the presence of a hermaphrodite flower in 50% of the inferior floscules and the presence of multiple embryo sacs of the Polygonum type within the same ovule in 15% of the pistils, are not related to low fertility. The majority of pollen grains are viable, in spite of the reduced number of pollen tubes within the style and ovary carpel, and a developing caryopsis was observed in 70% of self-pollinated pistils, indicating successful double fertilization from 2 days after anthesis (DAA). Nevertheless, abortion gradually increased from 2 until 7 DAA and remains elevated until 12 DAA, when caryopsis maturity is achieved. These data confirm low seed set in this accession and indicate that low fertility is not a consequence of abnormalities, either in the floral or gamete structures, or pollen tube rejection, but most likely a consequence of inbreeding depression.     

Detailed Characterization of Brain Dysfunction in a Long-Term Rodent Model of Critical Illness

Neurochemical Research - Critical illness encompasses a wide spectrum of life-threatening clinical conditions requiring intensive care. Our objective was to evaluate cognitive, inflammatory and...     

Experimental benznidazole treatment of Trypanosoma cruzi II strains isolated from children of the Jequitinhonha Valley,Minas Gerais,Brazil, with Chagas disease

Trypanosoma cruzi strains from distinct geographic areas show differences in drug resistance and association between parasites genetic and treatment response has been observed. Considering that benznidazole (BZ) can reduce the parasite burden and tissues damage, even in not cured animals and individuals, the goal is to assess the drug response to BZ of T. cruzi II strains isolated from children of the Jequitinhonha Valley, state of Minas Gerais, Brazil, before treatment. Mice infected and treated with BZ in both phases of infection were compared with the untreated and evaluated by fresh blood examination, haemoculture, polymerase chain reaction, conventional (ELISA) and non-conventional (FC-ALTA) serologies. In mice treated in the acute phase, a significant decrease in parasitaemia was observed for all strains. Positive parasitological and/or serological tests in animals treated during the acute and chronic (95.1-100%) phases showed that most of the strains were BZ resistant. However, beneficial effect was demonstrated because significant reduction (p < 0.05%) and/or suppression of parasitaemia was observed in mice infected with all strains (acute phase), associated to reduction/elimination of inflammation and fibrosis for two/eight strains. BZ offered some benefit, even in not cured animals, what suggest that BZ use may be recommended at least for recent chronic infection of the studied region.     

Differential Sensitivities of Fast- and Slow-Cycling Cancer Cells to Inosine Monophosphate Dehydrogenase 2 Inhibition by Mycophenolic Acid

As uncontrolled cell proliferation requires nucleotide biosynthesis, inhibiting enzymes that mediate nucleotide biosynthesis constitutes a rational approach to the management of oncological diseases. In practice, however, results of this strategy are mixed and thus elucidation of the mechanisms by which cancer cells evade the effect of nucleotide biosynthesis restriction is urgently needed. Here we explored the notion that intrinsic differences in cancer cell cycle velocity are important in the resistance toward inhibition of inosine monophosphate dehydrogenase (IMPDH) by mycophenolic acid (MPA). In short-term experiments, MPA treatment of fast-growing cancer cells effectively elicited G0/G1 arrest and provoked apoptosis, thus inhibiting cell proliferation and colony formation. Forced expression of a mutated IMPDH2, lacking a binding site for MPA but retaining enzymatic activity, resulted in complete resistance of cancer cells to MPA. In nude mice subcutaneously engrafted with HeLa cells, MPA moderately delayed tumor formation by inhibiting cell proliferation and inducing apoptosis. Importantly, we developed a lentiviral vector–based Tet-on label-retaining system that enables to identify, isolate and functionally characterize slow-cycling or so-called label-retaining cells (LRCs) in vitro and in vivo. We surprisingly found the presence of LRCs in fast-growing tumors. LRCs were superior in colony formation, tumor initiation and resistance to MPA as compared with fast-cycling cells. Thus, the slow-cycling compartment of cancer seems predominantly responsible for resistance to MPA.     

Differential expression of the pr1A gene in Metarhizium anisopliae and Metarhizium acridum across different culture conditions and during pathogenesis

The entomopathogenic fungi of the genus Metarhizium have several subtilisin-like proteases that are involved in pathogenesis and these have been used to investigate genes that are differentially expressed in response to different growth conditions. The identification and characterization of these proteases can provide insight into how the fungus is capable of infecting a wide variety of insects and adapt to different substrates. In addition, the pr1A gene has been used for the genetic improvement of strains used in pest control. In this study we used quantitative RT-PCR to assess the relative expression levels of the pr1A gene in M. anisopliae and M. acridum during growth in different culture conditions and during infection of the sugar cane borer, Diatraea saccharalis Fabricius. We also carried out a pathogenicity test to assess the virulence of both species against D. saccharalis and correlated the results with the pattern of pr1A gene expression. This analysis revealed that, in both species, the pr1A gene was differentially expressed under the growth conditions studied and during the pathogenic process. M. anisopliae showed higher expression of pr1A in all conditions examined, when compared to M. acridum. Furthermore, M. anisopliae showed a greater potential to control D. saccharalis. Taken together, our results suggest that these species have developed different strategies to adapt to different growing conditions.     

Reduced expression of SIRT1 is associated with diminished glucose-induced insulin secretion in islets from calorie-restricted rats

Alterations in food intake such as caloric restriction modulate the expression of SIRT1 and SIRT4 proteins that are involved in pancreatic β-cell function. Here, we search for a possible relationship between insulin secretion and the expression of SIRT1, SIRT4, PKC and PKA in islets from adult rats submitted to CR for 21 days. Rats were fed with an isocaloric diet (CTL) or received 60% (CR) of the food ingested by CTL. The dose-response curve of insulin secretion to glucose was shifted to the right in the CR compared with CTL islets (EC₅₀ of 15.1±0.17 and 10.5±0.11 mmol/L glucose). Insulin release by the depolarizing agents arginine and KCl was reduced in CR compared with CTL islets. Total islet insulin content and glucose oxidation were also reduced in CR islets. Leucine-stimulated secretion was similar in both groups, slightly reduced in CR islets stimulated by leucine plus glutamine but higher in CR islets stimulated by ketoisocaproate (KIC). Insulin secretion was also higher in CR islets stimulated by carbachol, compared with CTL islets. No differences in the rise of cytosolic Ca²⁺ concentrations stimulated by either glucose or KCl were observed between groups of islets. Finally, SIRT1, but not SIRT4, protein expression was lower in CR compared with CTL islets, whereas no differences in the expression of PKC and PKA proteins were observed. In conclusion, the lower insulin secretion in islets from CR rats was, at least in part, due to an imbalance between the expression of SIRT1 and SIRT4.     

Production and characterization of a collagenolytic serine proteinase by <Emphasis Type="Italic">Penicillium aurantiogriseum</Emphasis> URM 4622: A factorial study

A 2⁴ full factorial design was used to identify the main effects and interactions of the initial medium pH, soybean flour concentration, temperature and orbital agitation speed on extracellular collagenase production by Penicillium aurantiogriseum URM4622. The most significant variables for collagenase production were soybean flour concentration and initial medium pH that had positive main effects, and temperature that had a negative one. Protein concentration in soybean flour revealed to be a significant factor for the production of a collagenase serine proteinase. The most favorable production conditions were found to be 0.75% soybean flour, pH 8.0, 200 rpm, and 28°C, which led to a collagenase activity of 164 U. The enzyme showed an optimum activity at 37°C and pH 9.0, was stable over wide ranges of pH and temperature (6.0 ∼ 10.0 and 25 ∼ 45°C, respectively) and was strongly inhibited by 10 mM phenylmethylsulphonylfluoride. The firstorder rate constants for collagenase inactivation in the crude extract, calculated from semi-log plots of the residual activity versus time, were used in Arrhenius and Eyring plots to estimate the main thermodynamic parameters of thermoinactivation (E* _d = 107.4 kJ/mol and ΔH* _d = 104.7 kJ/mol). The enzyme is probably an extracellular neutral serine collagenase effective on azocoll, gelatin and collagen decomposition.