Mechanism of anticlastogenicity of Agaricus blazei Murill mushroom organic extracts in wild type CHO (K(1)) and repair deficient (xrs5) cells by chromosome aberration and sister chromatid exchange assays

Agaricus blazei Murill is a medicinal mushroom native to Brazil. The present work assessed the clastogenic and anticlastogenic potential of organic extracts (ethanol and chloroform/methanol) from the lineage AB97/11 in chinese hamster CHO-K(1) (wild type) and CHO-xrs5 (repair deficient) cells using the chromosome aberration (CA) and sister chromatid exchange (SCE) assays. In these experimental conditions were observed: (a) anticlastogenic effect at concentrations of 0.06 and 0.09% of the EtOH extract and at the 0.03 and 0.06% concentrations of the C/MetOH extract in CHO-K(1); (b) absence of protector effect on CHO-xrs5 cells; and (c) absence of protector effect in the SCE assay. These results indicate that organic extracts of A. blazei lineage AB97/11 present bio-antimutagenic type protective activity.     

Beta-aspartylpeptides as substrates of L-asparaginases from Escherichia coli and Erwinia chrysanthemi

L-Asparaginase is known to catalyze the hydrolysis of L-asparagine to L-aspartic and ammonia, but little is known about its action on peptides. When we incubated L-asparaginases purified either from Escherichia coli or Erwinia chrysanthemi - commonly used as chemotherapeutic agents because of their antitumour activity - with eight small beta-aspartylpeptides such as beta-aspartylserineamide, beta-aspartylalanineamide, beta-aspartylglycineamide and beta-aspartylglycine, we found that both L-asparaginases could catalyze the hydrolysis of five of them yielding L-aspartic acid and amino acids or peptides. Our data show that L-asparaginases can hydrolyze beta-aspartylpeptides and suggest that L-asparaginase therapy may affect the metabolism of beta-aspartylpeptides present in human body.     

Effects of crude extracts of Agaricus blazei on DNA damage and on rat liver carcinogenesis induced by diethylnitrosamine

The effects of crude extracts of the mushroom Agaricus blazei Murrill (Agaricaceae) on both DNA damage and placental form glutathione S-transferase (GST-P)-positive liver foci induced by diethylnitrosamine (DEN) were investigated. Six groups of adult male Wistar rats were used. For two weeks, animals of groups 3 to 6 were treated with three aqueous solutions of A. blazei (mean dry weight of solids being 1.2, 5.6, 11.5 and 11.5 mg/ml, respectively). After this period, groups 2 to 5 were given a single ip injection 200 mg/kg DEN and groups 1 and 6 were treated with 0.9% NaCl. All animals were subjected to 70% partial hepatectomy at week five and sacrificed 4, 24 and 48 h or 8 weeks after DEN or 0.9% NaCl treatments (10th week after the beginning of the experiment). The alkaline comet assay and GST-P-positive liver foci development were used to evaluate the influence of the mushroom extracts on liver cell DNA damage and on the initiation of liver carcinogenesis, respectively. Previous treatment with the highest concentration of A. blazei (11.5 mg/ml) significantly reduced DNA damage, indicating a protective effect against DEN-induced liver cytotoxicity/genotoxicity. However, the same dose of mushroom extract significantly increased the number of GST-P-positive liver foci.     

Biomanipulation as an Application of Food-Chain Theory: Constraints, Synthesis, and Recommendations for Temperate Lakes

The aim of this review is to identify problems, find general patterns, and extract recommendations for successful biomanipulation. An important conclusion is that the pelagic food chain from fish to algae may not be the only process affected by a biomanipulation. Instead, this process should be viewed as the “trigger” for secondary processes, such as establishment of submerged macrophytes, reduced internal loading of nutrients, and reduced resuspension of particles from the sediment. However, fish reduction also leads to a high recruitment of young-of-the-year (YOY) fish, which feed extensively on zooplankton. This expansion of YOY the first years after fish reduction is probably a major reason for less successful biomanipulations. Recent, large-scale biomanipulations have made it possible to update earlier recommendations regarding when, where, and how biomanipulation should be performed. More applicable recommendations include (1) the reduction in the biomass of planktivorous fish should be 75% or more; (2) the fish reduction should be performed efficiently and rapidly (within 1–3 years); (3) efforts should be made to reduce the number of benthic feeding fish; (4) the recruitment of YOY fish should be reduced; (5) the conditions for establishment of submerged macrophytes should be improved; and (6) the external input of nutrients (phosphorus and nitrogen) should be reduced as much as possible before the biomanipulation. Recent biomanipulations have shown that, correctly performed, the method also achieves results in large, relatively deep and eutrophic lakes, at least in a 5-year perspective. Although repeated measures may be necessary, the general conclusion is that biomanipulation is not only possible, but also a relatively inexpensive and attractive method for management of eutrophic lakes, and in particular as a follow-up measure to reduced nutrient load. Received 14 April 1998; accepted 31 August 1998     

CDKN3 mRNA as a Biomarker for Survival and Therapeutic Target in Cervical Cancer

The cyclin-dependent kinase inhibitor 3 (CDKN3) gene, involved in mitosis, is upregulated in cervical cancer (CC). We investigated CDKN3 mRNA as a survival biomarker and potential therapeutic target for CC. CDKN3 mRNA was measured in 134 CC and 25 controls by quantitative PCR. A 5-year survival study was conducted in 121 of these CC patients. Furthermore, CDKN3-specific siRNAs were used to investigate whether CDKN3 is involved in proliferation, migration, and invasion in CC-derived cell lines (SiHa, CaSki, HeLa). CDKN3 mRNA was on average 6.4-fold higher in tumors than in controls (p = 8 x 10⁻⁶, Mann-Whitney). A total of 68.2% of CC patients over expressing CDKN3 gene (fold change ≥ 17) died within two years of diagnosis, independent of the clinical stage and HPV type (Hazard Ratio = 5.0, 95% CI: 2.5–10, p = 3.3 x 10⁻⁶, Cox proportional-hazards regression). In contrast, only 19.2% of the patients with lower CDKN3 expression died in the same period. In vitro inactivation of CDKN3 decreased cell proliferation on average 67%, although it had no effect on cell migration and invasion. CDKN3 mRNA may be a good survival biomarker and potential therapeutic target in CC.     

Utilization of seagrass habitats by juvenile groupers and snappers in Banten Bay,Banten Province,Indonesia

Coastal development in Banten Bay, Indonesia, decreased seagrass coverage to only 1.5% of its surface area. We investigated the importance of seagrass as habitat for juvenile groupers (Serranidae) and snappers (Lutjanidae), by performing beam trawl hauls on a weekly basis in two seagrass locations and one mudflat area, and monthly trawl hauls in three different microhabitats (dense, mixed and patchy seagrass) in one of the seagrass locations. We studied the effects of location and microhabitat, as well as temporal patterns (diel, weekly and monthly) on the probability of occurrence and abundance of the most abundant grouper (Orange-spotted grouper, Epinephelus coioides) and snapper (Russell’s snapper, Lutjanus russellii). We found that both species were almost exclusively found in seagrass locations, with a preference for microhabitats of high complexity (dense and mixed microhabitats). L. russellii had a higher probability of catch and abundance during the night, most probably because of its ability to avoid the beam trawl during daytime sampling. In addition there was an effect of week and month on the presence and abundance of both species, but patterns were unclear, probably because of high fishing pressure on juvenile groupers and snappers by push net fishermen. Groupers and snappers mainly fed on abundant shrimps, and to a lesser extent on fish. Moreover, juveniles find protection against predators in seagrass, which confirmed the critical role of quantity and quality of seagrass areas for juvenile groupers and snappers in Banten Bay.     

Cryopreservation at −80°C of Agaricus blazei on rice grains

The preservation of Agaricus blazei is generally done by mycelial subculturing, but this technique may cause genetic degenerations. Despite this, there is not an efficient protocol established to preserve this fungus and cryopreservation could be an alternative. This study aimed to evaluate two freezing protocols for cryopreservation at −80°C of A. blazei strains. Five fungus strains grown on rice grains with husk and were transferred to glycerol (10%) in cryovials. Next, the cryovials were submitted to two freezing temperature protocols: (1) cryopreservation starting at 25°C, then at 8°C for 30 min and kept at −80°C; (2) cryopreservation starting at 25°C, then 8°C for 30 min, −196°C for 15 min and kept at −80°C. After 1 year of cryopreservation, the cryovials were thawed in a water bath at 30°C for 15 min and transferred to malt extract agar medium. It was concluded that the one-year cryopreservation process of A. blazei, grown on rice grains and cryopreserved at −80°C in glycerol 10%, is viable. The slow freezing, from 8 to −80°C, is effective whereas the fast freezing, from 8 to −196°C and then to −80°C, is ineffective. The different genetic characteristics among the strains of this fungus do not interfere in the cryopreservation process.     

Photocycle of the flavin-binding photoreceptor AppA, a bacterial transcriptional antirepressor of photosynthesis genes

The flavoprotein AppA from Rhodobacter sphaeroides contains an N-terminal domain belonging to a new class of photoreceptors designated BLUF domains. AppA was shown to control photosynthesis gene expression in response to blue light and oxygen tension. We have investigated the photocycle of the AppA BLUF domain by ultrafast fluorescence, femtosecond transient absorption, and nanosecond flash-photolysis spectroscopy. Time-resolved fluorescence experiments revealed four components of flavin adenine dinucleotide (FAD) excited-state decay, with lifetimes of 25 ps, 150 ps, 670 ps, and 3.8 ns. Ultrafast transient absorption spectroscopy revealed rapid internal conversion and vibrational cooling processes on excited FAD with time constants of 250 fs and 1.2 ps, and a multiexponential decay with effective time constants of 90 ps, 590 ps, and 2.7 ns. Concomitant with the decay of excited FAD, the rise of a species with a narrow absorption difference band near 495 nm was detected which spectrally resembles the long-living signaling state of AppA. Consistent with these results, the nanosecond flash-photolysis measurements indicated that formation of the signaling state was complete within the time resolution of 10 ns. No further changes were detected up to 15 micros. The quantum yield of the signaling-state formation was determined to be 24%. Thus, the signaling state of the AppA BLUF domain is formed on the ultrafast time scale directly from the FAD singlet excited state, without any apparent intermediate, and remains stable over 12 decades of time. In parallel with the signaling state, the FAD triplet state is formed from the FAD singlet excited state at 9% efficiency as a side reaction of the AppA photocycle.