Evaluating the Effects of Separate and Concomitant Use of MK-2206 and Salinomycin on Prostate Cancer Cell Line

Background:Prostate cancer is known as one of the most prevalent health disorders in the male population globally. The aim of the current study was to evaluate the effects of separate and concomitant use of MK-2206 and salinomycin on prostate cancer cell line.Methods:The antitumor potential of separate and concomitant use of MK-2206 and salinomycin was evaluated in a panel of prostate cancer cell line (PC-3). To get insights into the underlying mechanism of action, different assays including the rate of apoptosis, cell viability, and gene expression were performed in treated prostate cancer cells.Results:A significant reduction was detected in the viability percentage of prostate cancer cells (p< 0.001) and the rate of Akt expression (p< 0.001) in all salinomycin, MK-2206, and salinomycin+MK-2206 groups compared to the negative control group. Furthermore, in comparison with the negative control group, there was a notable increase in both the rate of Bad expression (p< 0.001) and prostate cancer cells apoptosis after salinomycin, MK-2206, and salinomycin+MK-2206 treatments. Moreover, the concomitant use of salinomycin+MK-2206 revealed synergistic improvements regarding the viability of prostate cancer cells and the rate of the Akt and Bad expressions compared to the separate administration of salinomycin and MK-2206 (all p< 0.05)Conclusion:The findings of the present study may contribute to improving the efficacy of the therapies regarding the management of prostate cancer and providing a beneficial strategy in clinical trials.Key Words: Apoptosis, Gene Expression, MK 2206, Prostatic Neoplasms, Salinomycin     

Detection of Nitrosomonas spp. by polymerase chain reaction

Abstract A unique genomic DNA fragment was isolated from Nitrosomonas europaea ATCC 19718. Based on the sequence of this fragment, oligonucleotide primers for polymerase chain reaction amplification were prepared which amplify sequences of 775 and 658 bp. The predicted DNA fragments were both amplified from the genome of N. europaea and a Nitrosomonas spp. isolated from a local oxidation pond. The primers failed to amplify DNA from the genomes of the ammonia oxidiser Nitrosolobous multiformis , the nitrite oxidiser Nitrococcus mobilis as well as from the genomes of other unrelated heterotrophic bacteria. These DNA sequences could be amplified from 0.01 ng of N. europaea genomic DNA or from 100 intact cells, and it was possible to detect Nitrosomonas DNA in a DNA mixture extracted from water samples drawn from a local oxidation pond.     

Heterogeneous effects of cholecystokinin on neuronal response properties in deep layers of rat barrel cortex

Abstract

Cholecystokinin (CCK) is one of the most studied neuropeptides in the brain. In this study, we investigated the effects of CCK-8s and LY225910 (CCK₂ receptor antagonist) on properties of neuronal response to natural stimuli (whisker deflection) in deep layers of rat barrel cortex. This study was done on 20 male Wistar rats, weighing 230–260?g. CCK-8s (300?nmol/rat) and LY225910 (1?µmol/rat) were administered intracerebroventricularly (ICV). Neuronal responses to deflection of principal (PW) and adjacent (AW) whiskers were recorded in the barrel cortex using tungsten microelectrodes. Computer controlled mechanical displacement was used to deflect whiskers individually or in combination at 30?ms inter-stimulus intervals. ON and OFF responses for PW and AW deflections were measured. A condition-test ratio (CTR) was computed to quantify neuronal responses to whisker interaction. ICV administration of CCK-8s and LY225910 had heterogeneous effects on neuronal spontaneous activity, ON and OFF responses to PW and/or AW deflections, and CTR for both ON and OFF responses. The results of this study demonstrated that CCK-8s can modulate neuronal response properties in deep layers of rat barrel cortex probably via CCK₂ receptors.     

Genistein effects on growth and cell cycle ofCandida albicans

Microbial virulence is generally considered to be multifactorial with infection resulting from the sum of several globally regulated virulence factors. Estrogen may serve as a signal for global virulence induction in Candida albicans. Nonsteroidal estrogens and estrogen receptor antagonists may therefore have interesting effects on yeast and their virulence factors. Growth of C. albicans was monitored by viable plate counts at timed intervals after inoculation into yeast nitrogen broth plus glucose. To determine if increased growth of yeast in the presence of estradiol was due to tyrosine kinase-mediated signaling, we measured growth in the presence of genistein, estradiol or genistein plus estradiol and compared these conditions to controls, which were not supplemented with either compound. Unexpectedly, genistein stimulated growth of C. albicans. In addition, genistein was found to increase the rate of germination (possibly reflecting release from G(0) into G(1) cell cycle phase) and also increased Hsp90 expression, demonstrated by a dot blot technique which employed a commercial primary antibody detected with chemiluminescence with horseradish peroxidase-labeled secondary antibody. These biological effects may be attributable to genistein's activity as a phytoestrogen. In contrast, nafoxidine suppressed growth of Candida and mildly diminished Hsp90 expression. This study raises the possibility of receptor cross-talk between estrogen and isoflavinoid compounds, and antiestrogens which may affect the same signaling system, though separate targets for each compound were not ruled out.     

Network resonance can be generated independently at distinct levels of neuronal organization

Resonance is defined as maximal response of a system to periodic inputs in a limited frequency band. Resonance may serve to optimize inter-neuronal communication, and has been observed at multiple levels of neuronal organization. However, it is unknown how neuronal resonance observed at the network level is generated and how network resonance depends on the properties of the network building blocks. Here, we first develop a metric for quantifying spike timing resonance in the presence of background noise, extending the notion of spiking resonance for in vivo experiments. Using conductance-based models, we find that network resonance can be inherited from resonances at other levels of organization, or be intrinsically generated by combining mechanisms across distinct levels. Resonance of membrane potential fluctuations, postsynaptic potentials, and single neuron spiking can each be generated independently of resonance at any other level and be propagated to the network level. At all levels of organization, interactions between processes that give rise to low- and high-pass filters generate the observed resonance. Intrinsic network resonance can be generated by the combination of filters belonging to different levels of organization. Inhibition-induced network resonance can emerge by inheritance from resonance of membrane potential fluctuations, and be sharpened by presynaptic high-pass filtering. Our results demonstrate a multiplicity of qualitatively different mechanisms that can generate resonance in neuronal systems, and provide analysis tools and a conceptual framework for the mechanistic investigation of network resonance in terms of circuit components, across levels of neuronal organization.     

Brain Cytochrome Oxidase in Alzheimer''s Disease

A recent demonstration of markedly reduced (-50%) activity of cytochrome oxidase (CO; complex 4), the terminal enzyme of the mitochondrial enzyme transport chain, in platelets of patients with Alzheimer's disease (AD) suggested the possibility of a systemic and etiologically fundamental CO defect in AD. To determine whether a CO deficiency occurs in AD brain, we measured the activity of CO in homogenates of autopsied brain regions of 19 patients with AD and 30 controls matched with respect to age, postmortem time, sex, and, as indices of agonal status, brain pH and lactic acid concentration. Mean CO activity in AD brain was reduced in frontal (-26%: p less than 0.01), temporal (-17%; p less than 0.05), and parietal (-16%; not significant, p = 0.055) cortices. In occipital cortex and putamen, mean CO levels were normal, whereas in hippocampus, CO activity, on average, was nonsignificantly elevated (20%). The reduction of CO activity, which is tightly coupled to neuronal metabolic activity, could be explained by hypofunction of neurons, neuronal or mitochondrial loss, or possibly by a more primary, but region-specific, defect in the enzyme itself. The absence of a CO activity reduction in all of the examined brain areas does not support the notion of a generalized brain CO abnormality. Although the functional significance of a 16-26% cerebral cortical CO deficit in human brain is not known, a deficiency of this key energy-metabolizing enzyme could reduce energy stores and thereby contribute to the brain dysfunction and neurodegenerative processes in AD.     

Hydroxyl distribution in sugar structure and its contributory role in the inhibition of Stachybotrys microspora β-glucosidase (bglG)

Stachybotrys microspora is a filamentous fungus that produces various β-glucosidases, of which two have already been characterized. The present study reports on the production of a third one, named bglG, in the presence of d-glucose used as a sole carbon source, and on its subsequent purification and characterization. Although efficiently produced in the presence of d-glucose, bglG continues to be highly inhibited by this sugar. In fact, the addition of d-glucose significantly decreases the glucose formation rates during the hydrolysis of pNPG. This work reports on the effect of various carbohydrates on bglG activity in order to understand the mechanisms adopted by d-glucose to inhibit this enzyme. The findings indicate that bglG is strongly inhibited by d-glucose (44% of the relative activity at 5 mM), d-glucitol (96% of the relative activity), d-mannose (56% of the relative activity), cellobiose and maltose (72% and 71% of the relative activity, respectively). On the other hand, d-galactose, d-fructose, lactose, and sucrose have no effect on bglG activity. Similarly, several isomers, such as 2-acetamido-2-deoxy-d-glucose and 2-deoxy-d-arabino-hexose (2-deoxy-d-glucose) were noted to bring no change on the relative activity of bglG. d-xylose and xylitol, on the other hand, enhanced bglG activity up to 123% and 120% of relative activity, respectively. Accordingly, the configuration, epimerisation, isomerisation, and substitutions played key roles in bglG inhibition. The effect of the combination of iron (the best activator of bglG, 161%) with some of those additives was also investigated. The findings revealed that, while a combination of iron at a concentration of 10 mM with d-glucose resulted in a two-fold decrease in bglG inhibition (84% at 5 mM), iron maintained the same effect with the remainder of the additives being tested.     

Molecular detection of nematicidal crystalliferous Bacillus thuringiensis strains of Iran and evaluation of their toxicity on free-living and plant-parasitic nematodes

The characterization of nematode-effective strains and cry genes in the Iranian Bacillus thuringiensis (Bt) collection (70 isolates) is presented. Characterization was based on PCR analysis using 12 specific primers for cry5, cry6, cry12, cry13, cry14, and cry21 genes encoding proteins active against nematodes, crystal morphology, and protein band patterns as well as their nematicidal activity on root-knot nematode (Meloidogyne incognita) and two free-living nematodes (Chiloplacus tenuis and Acrobeloides enoplus). PCR results with primers for these genes showed that 22 isolates (31.5%) contain a minimum of one nematode-active cry gene. Strains containing the cry6 gene were the most abundant and represent 22.8% of the isolates. Bt strains harboring cry14 genes were also abundant (14.2%). cry21 and cry5 genes were less abundant, found in 4.2% and 2.8% of the strains, respectively. In total, six different nematode-active cry gene profiles were detected in this collection. Four isolates did not show the expected PCR product size for cry5, cry6, and cry21 genes; they might contain potentially novel insecticidal crystal protein genes. Twenty-two Bt isolates containing nematode-active cry genes were selected for preliminary bioassays on M. incognita. Based on these bioassays, four isolates were selected for detailed bioassays. Isolates YD5 and KON4 at 2 x 10(8) CFU/mL concentrations showed 77% and 81% toxicity on M. incognita, respectively. The free-living nematodes C. tenuis and A. enoplus were more susceptible and the highest mortality was observed within 48 h of incubation at all of the concentrations tested. Maximum mortality was recorded for isolates SN1 and KON4 at 2 x 10(8) CFU/mL concentrations and resulted in 68% and 77% adults deaths of C. tenuis and 68% and 72% for A. enoplus, respectively. Our results showed that PCR is a useful technique for toxicity prediction of nematicidal Bt isolates.     

Comparison of dynamic responses of cellular metabolites in <Emphasis Type=Italic>Escherichia coli</Emphasis> to pulse addition of substrates

We conducted an integrated study of cell growth parameters, product formation, and the dynamics of intracellular metabolite concentrations using Escherichia coli with genes knocked out in the glycolytic and oxidative pentose phosphate pathway (PPP) for glucose catabolism. We investigated the same characteristics in the wild-type strain, using acetate or pyruvate as the sole carbon source. Dramatic effects on growth parameters and extracellular and intracellular metabolite concentrations were observed after blocking either glycolytic breakdown of glucose by inactivation of phosphoglucose isomerase (disruption of pgi gene) or pentose phosphate breakdown of glucose by inactivation of glucose-6-phosphate dehydrogenase (disruption of zwf gene). Reducing power (NADPH) was mainly produced through PPP when the pgi gene was knocked out, while NADPH was produced through the tricarboxylic acid (TCA) cycle by isocitrate dehydrogenase or NADP-linked malic enzyme when the zwf gene was knocked out. As expected, when the pgi gene was knocked out, intracellular concentrations of PPP metabolites were high and glycolytic and concentrations of TCA cycle pathway metabolites were low. In the zwf gene knockout, concentrations of PPP metabolites were low and concentrations of intracellular glycolytic and TCA cycle metabolites were high.