Increase in therapeutic index of doxorubicin and vinblastine by aptameric oligonucleotide in human T lymphoblastic drug-sensitive and multidrug-resistant cells

Aptameric GT oligomers are a new class of potential anticancer molecules that inhibit the growth of human cancer cell lines by binding to specific nuclear proteins. We demonstrated that an aptameric GT oligonucleotide increased the therapeutic index of doxorubicin and vinblastine in T lymphoblastic drug-sensitive and multidrug-resistant (MDR) cells. The doxorubicin ID50 decreased 6.5-fold by coadministration of 1 microM GT to CCRF-CEM cells and by 24-fold by coadministration of 0.75 microM GT to CEM-VLB300 cells. In CEM-VLB300 cells, the vinblastine ID50 decreased 11-fold by coadministration of 0.5 microM GT. Control CT sequence did not potentiate the drugs in either CCRF-CEM or CEM-VLB300 cells. The ability of GT to bind to specific nuclear proteins in cancer cells related to the increase in the therapeutic index of doxorubicin and vinblastine. No cooperation was detected by the administration of GT oligomer together with doxorubicin to rat differentiated thyroid FRTL-5 cells and to normal human lymphocytes. These cells did not show binding of GT to the specific nuclear proteins, and they were not sensitive to the cytotoxic action of the GT sequence. Drug potentiation by GT not involving normal human lymphocytes might be exploited to develop a more selective treatment of drug-sensitive and MDR tumors.     

Studying Memory Encoding to Promote Reliable Engagement of the Medial Temporal Lobe at the Single-Subject Level

The medial temporal lobe (MTL)—comprising hippocampus and the surrounding neocortical regions—is a targeted brain area sensitive to several neurological diseases. Although functional magnetic resonance imaging (fMRI) has been widely used to assess brain functional abnormalities, detecting MTL activation has been technically challenging. The aim of our study was to provide an fMRI paradigm that reliably activates MTL regions at the individual level, thus providing a useful tool for future research in clinical memory-related studies. Twenty young healthy adults underwent an event-related fMRI study consisting of three encoding conditions: word-pairs, face-name associations and complex visual scenes. A region-of-interest analysis at the individual level comparing novel and repeated stimuli independently for each task was performed. The results of this analysis yielded activations in the hippocampal and parahippocampal regions in most of the participants. Specifically, 95% and 100% of participants showed significant activations in the left hippocampus during the face-name encoding and in the right parahippocampus, respectively, during scene encoding. Additionally, a whole brain analysis, also comparing novel versus repeated stimuli at the group level, showed mainly left frontal activation during the word task. In this group analysis, the face-name association engaged the HP and fusiform gyri bilaterally, along with the left inferior frontal gyrus, and the complex visual scenes activated mainly the parahippocampus and hippocampus bilaterally. In sum, our task design represents a rapid and reliable manner to study and explore MTL activity at the individual level, thus providing a useful tool for future research in clinical memory-related fMRI studies.     

Determination of Parameters for the Supercritical Extraction of Antioxidant Compounds from Green Propolis Using Carbon Dioxide and Ethanol as Co-Solvent

The aim of this study was to determine the best processing conditions to extract Brazilian green propolis using a supercritical extraction technology. For this purpose, the influence of different parameters was evaluated such as S/F (solvent mass in relation to solute mass), percentage of co-solvent (1 and 2% ethanol), temperature (40 and 50°C) and pressure (250, 350 and 400 bar) using supercritical carbon dioxide. The Global Yield Isotherms (GYIs) were obtained through the evaluation of the yield, and the chemical composition of the extracts was also obtained in relation to the total phenolic compounds, flavonoids, antioxidant activity and 3,5-diprenyl-4-hydroxicinnamic acid (Artepillin C) and acid 4-hydroxycinnamic (p-coumaric acid). The best results were identified at 50°C, 350 bar, 1% ethanol (co-solvent) and S/F of 110. These conditions, a content of 8.93±0.01 and 0.40±0.05 g/100 g of Artepillin C and p-coumaric acid, respectively, were identified indicating the efficiency of the extraction process. Despite of low yield of the process, the extracts obtained had high contents of relevant compounds, proving the viability of the process to obtain green propolis extracts with important biological applications due to the extracts composition.     

SIRT5 regulation of ammonia-induced autophagy and mitophagy

In liver the mitochondrial sirtuin, SIRT5, controls ammonia detoxification by regulating CPS1, the first enzyme of the urea cycle. However, while SIRT5 is ubiquitously expressed, urea cycle and CPS1 are only present in the liver and, to a minor extent, in the kidney. To address the possibility that SIRT5 is involved in ammonia production also in nonliver cells, clones of human breast cancer cell lines MDA-MB-231 and mouse myoblast C2C12, overexpressing or silenced for SIRT5 were produced. Our results show that ammonia production increased in SIRT5-silenced and decreased in SIRT5-overexpressing cells. We also obtained the same ammonia increase when using a new specific inhibitor of SIRT5 called MC3482. SIRT5 regulates ammonia production by controlling glutamine metabolism. In fact, in the mitochondria, glutamine is transformed in glutamate by the enzyme glutaminase, a reaction producing ammonia. We found that SIRT5 and glutaminase coimmunoprecipitated and that SIRT5 inhibition resulted in an increased succinylation of glutaminase. We next determined that autophagy and mitophagy were increased by ammonia by measuring autophagic proteolysis of long-lived proteins, increase of autophagy markers MAP1LC3B, GABARAP, and GABARAPL2, mitophagy markers BNIP3 and the PINK1-PARK2 system as well as mitochondrial morphology and dynamics. We observed that autophagy and mitophagy increased in SIRT5-silenced cells and in WT cells treated with MC3482 and decreased in SIRT5-overexpressing cells. Moreover, glutaminase inhibition or glutamine withdrawal completely prevented autophagy. In conclusion we propose that the role of SIRT5 in nonliver cells is to regulate ammonia production and ammonia-induced autophagy by regulating glutamine metabolism.     

Analysis of the Role of Interleukin 6 Receptor Haplotypes in the Regulation of Circulating Levels of Inflammatory Biomarkers and Risk of Coronary Heart Disease

Variants at the interleukin 6 receptor (IL6R) gene regulate inflammation and are associated with risk of coronary heart disease (CHD). The aim of the present study was to investigate the effects of IL6R haplotypes on circulating levels of inflammatory biomarkers and risk of CHD. We performed a discovery analysis in SHEEP, a myocardial infarction (MI) case control study (n = 2,774) and replicated our results in two large, independent European populations, PROCARDIS, a CHD case control study (n = 7,998), and IMPROVE (n = 3,711) a prospective cardiovascular cohort study. Two major haplotype blocks (rs12083537A/G and rs4075015A/T—block 1; and rs8192282G/A, rs4553185T/C, rs8192284A/C, rs4240872T/C and rs7514452T/C—block 2) were identified in the IL6R gene. IL6R haplotype associations with C-reactive protein (CRP), fibrinogen, IL6, soluble IL6R (sIL6R), IL6, IL8 and TNF-α in SHEEP, CRP and fibrinogen in PROCARDIS and CRP in IMPROVE as well as association with risk of MI and CHD, were analyzed by THESIAS. Haplotypes in block 1 were associated neither with circulating inflammatory biomarkers nor with the MI/CHD risk. Haplotypes in block 2 were associated with circulating levels of CRP, in all three study populations, with fibrinogen in SHEEP and PROCARDIS, with IL8 and sIL6Rin SHEEP and with a modest, non significant, increase (7%) in MI/CHD risk in the three populations studied. Our results indicate that IL6R haplotypes regulate the circulating levels of inflammatory biomarkers. Lack of association with the risk of CHD may be explained by the combined effect of SNPs with opposite effect on the CHD risk, the sample size as well as by structural changes affecting sIL6R stability in the circulation.     

Effortless Attention as a Biomarker for Experienced Mindfulness Practitioners

Objective

The present study aimed at comparing frontal beta power between long-term (LTM) and first-time meditators (FTM), before, during and after a meditation session. We hypothesized that LTM would present lower beta power than FTM due to lower effort of attention and awareness.

Methods

Twenty one participants were recruited, eleven of whom were long-term meditators. The subjects were asked to rest for 4 minutes before and after open monitoring (OM) meditation (40 minutes).

Results

The two-way ANOVA revealed an interaction between the group and moment factors for the Fp1 (p<0.01), F7 (p = 0.01), F3 (p<0.01), Fz (p<0.01), F4 (p<0.01), F8 (p<0.01) electrodes.

Conclusion

We found low power frontal beta activity for LTM during the task and this may be associated with the fact that OM is related to bottom-up pathways that are not present in FTM.

Significance

We hypothesized that the frontal beta power pattern may be a biomarker for LTM. It may also be related to improving an attentive state and to the efficiency of cognitive functions, as well as to the long-term experience with meditation (i.e., life-time experience and frequency of practice).     

Direct Visualization of the Action of Triton X-100 on Giant Vesicles of Erythrocyte Membrane Lipids

The raft hypothesis proposes that microdomains enriched in sphingolipids, cholesterol, and specific proteins are transiently formed to accomplish important cellular tasks. Equivocally, detergent-resistant membranes were initially assumed to be identical to membrane rafts, because of similarities between their compositions. In fact, the impact of detergents in membrane organization is still controversial. Here, we use phase contrast and fluorescence microscopy to observe giant unilamellar vesicles (GUVs) made of erythrocyte membrane lipids (erythro-GUVs) when exposed to the detergent Triton X-100 (TX-100). We clearly show that TX-100 has a restructuring action on biomembranes. Contact with TX-100 readily induces domain formation on the previously homogeneous membrane of erythro-GUVs at physiological and room temperatures. The shape and dynamics of the formed domains point to liquid-ordered/liquid-disordered (Lo/Ld) phase separation, typically found in raft-like ternary lipid mixtures. The Ld domains are then separated from the original vesicle and completely solubilized by TX-100. The insoluble vesicle left, in the Lo phase, represents around 2/3 of the original vesicle surface at room temperature and decreases to almost 1/2 at physiological temperature. This chain of events could be entirely reproduced with biomimetic GUVs of a simple ternary lipid mixture, 2:1:2 POPC/SM/chol (phosphatidylcholine/sphyngomyelin/cholesterol), showing that this behavior will arise because of fundamental physicochemical properties of simple lipid mixtures. This work provides direct visualization of TX-100-induced domain formation followed by selective (Ld phase) solubilization in a model system with a complex biological lipid composition.     

Vocal Repertoire of Golden-backed Uakaris (<Emphasis Type="Italic">Cacajao melanocephalus</Emphasis>): Call Structure and Context

The study of vocal behavior can reveal important aspects of how and why a species communicates in relation to ecological and social challenges. We here focus on vocal communication in golden-backed uakaris (Cacajao melanocephalus), diurnal, pitheciine monkeys that exhibit fission-fusion social organization and typically inhabit dense forests that limit the potential for visual communication. Moreover, the species spends little time engaged in tactile or olfactory communication, e.g., social grooming and scent marking, respectively. Hence, vocalizations may be very important for the coordination of social organization in these monkeys. We 1) categorized golden-backed uakari vocalizations, 2) ascertained their behavioral context, and 3) investigated whether golden-backed uakari calls can encode information about the signaler. We observed the monkeys during 2 wet seasons in the flooded igapó forest of Jaú National Park, Brazil. We showed that golden-backed uakaris have 9 call types in their vocal repertoire, all distinguishable by ear and from analysis of spectrograms. Some calls, e.g., play-specific calls, were used only in particular behavioral contexts, and by individuals of specific age, whereas others were emitted under a range of situations. The structure of the loud tchó call varied among individuals, and according to behavioral context, i.e., whether individuals were foraging/feeding, traveling, or performing agonistic interactions. This knowledge of the species’ vocal repertoire is valuable for surveying the monkeys acoustically in habitats where visual surveys are difficult.     

Flow-regulated glucose and lipid metabolism in adipose tissue,endothelial cell and hepatocyte cultures in a modular bioreactor

Static cell culture has serious limitations in its ability to represent cellular behaviour within a live organism. In vivo, cells are constantly exposed to the flow of bodily fluids and contact with other cell types. Bioreactors provide the opportunity to study cells in an environment that more closely resembles the in vivo setting because cell cultures can be exposed to dynamic flow in contact with or in proximity to other cell types. In this study we compared the metabolic profile of a dynamic cell culture system to that of a static cell culture in three different cellular phenotypes: adipocytes, endothelial cells and hepatocytes. Albumin, glucose, free fatty acids, glycerol, and lactate were measured over 48 h. We show that all three cell types have increased glucose uptake in the presence of flow; lactate release was also significantly affected. We provide robust evidence that the presence of flow significantly modifies cellular metabolism. While flow provides a more uniform nutrient distribution and increases metabolite turnover, our results indicate that different cell types have specific metabolic responses to flow, suggesting cell-specific flow-regulated activation of metabolite signalling pathways.