Effects of chlorogenic acid and its metabolites on spontaneous locomotor activity in mice

Chlorogenic acid possessed a weak caffeine-like psychostimulant property when assessed for its effect on spontaneous locomotor activity in mice. In the evaluation of the effects for the major metabolites of chlorogenic acid which were detected upon incubation with rat feces and/or excreted in urine after oral administration to rats, caffeic and m-coumaric acids were found to be the principal active metabolites, while the others contributed little to this caffeine-like psychostimulant activity.     

An interaction between benzodiazepines and neuroactive steroids at GABA A receptors in cultured hippocampal neurons

Neurosteroids are modulators of several receptors and ion channels and are implicated in the pathophysiology of several neuropsychiatric diseases including hepatic encephalopathy (HE). The neurosteroid, allopregnanolone, a positive allosteric modulator of GABA_A receptors, accumulates in the brains of HE patients where it can potentiate GABA_A receptor-mediated responses. Attenuation of the effects of neurosteroids on GABA-ergic neurotransmission is therefore of interest for the management of HE. In the present study, we determined the effect of the benzodiazepine partial inverse agonist, Ro15-4513, and the benzodiazepine antagonist, flumazenil on modulation of the GABA_A mediated chloride currents by allopregnanolone and on spontaneous synaptic activity in cultured hippocampal neurons using the patch-clamp technique. Allopregnanolone (0.03–0.3 μM), dose-dependently potentiated GABA-induced currents, an action significantly reduced by Ro15-4513 (10 μM). In contrast, flumazenil (10 μM) had no effect on the ability of allopregnanolone to potentiate GABA_A currents but it blocked the effects of Ro15-4513. The frequency of spontaneous synaptic activity was significantly reduced in the presence of allopregnanolone (0.1 μM) from 1.5 ± 0.7 to 0.1 ± 0.04 Hz. This action was partially reversed by Ro15-4513 (10 μM) but was not significantly influenced by flumazenil (10 μM). These findings suggest that the beneficial affects of Ro15-4513 in experimental HE result from attenuation of the effects of neurosteroids at GABA_A receptors. Our results may provide a rational basis for the use of benzodiazepine inverse agonists in the management and treatment of hepatic encephalopathy in patients with liver failure.     

Development of glutathione-coupled cantilever for the single-molecule force measurement by scanning force microscopy

The accuracy and the fidelity of a single-molecule force measurement largely rely on how the molecule of interest is attached to the solid substrate surface (bead, cantilever, cover glass and etc.). A site-specific attachment of a protein without affecting its structure and enzymatic function has been a major concern. Here, we established a glutathione-coupled cantilever to which any glutathione S-transferase (GST)-fused proteins can be attached in a desired direction. The rupture force between glutathione and GST was approximately 100 pN on average. By using this cantilever, we succeeded in measuring the interaction force between importin alpha and importin beta.     

Antioxidant activity of flavonoids evaluated with myoglobin method

Antioxidant activities of four flavonoids (rutin, quercetin, luteolin, and kaempferol) and two non-flavonoids (chlorogenic acid and pyrocatechol) against four reactive oxygen species (ROS) have been measured with a myoglobin method developed by our group. The myoglobin method uses the absorbance changes of myoglobin (a probe molecule) due to the reaction with the ROS as an indicator for the antioxidant activity measurement. Myoglobin protective ratio (MPR) was defined to express the antioxidant activities of the specimens. Antioxidant activities against hypochlorite ion, hydroxyl radical, peroxyl radical, and peroxynitrite were measured with the myoglobin method. The antioxidant activities were comprehensively evaluated by plotting MPR against four ROS and vitamin C equivalent concentration evaluated by DPPH quenching method in 5-axe cobweb charts. The four flavonoids show a very similar pattern in the 5-axe cobweb charts, while the patterns of two non-flavonoids are quite different from that of the flavonoids. This procedure combining the myoglobin method with the cobweb charts is useful in the evaluation of antioxidant activities of plant-derived food, and also can be extended to monitor antioxidant condition of media for plant cell cultures.     

Hypothesis with abnormal amino acid metabolism in depression and stress vulnerability in Wistar Kyoto rats

While abnormalities in monoamine metabolism have been investigated heavily per potential roles in the mechanisms of depression, the contribution of amino acid metabolism in the brain remains not well understood. In additional, roles of the hypothalamus–pituitary–adrenal axis in stress-regulation mechanisms have been of much focus, while the contribution of central amino acid metabolism to these mechanisms has not been well appreciated. Therefore, whether depression-like states affect amino acid metabolism and their potential roles on stress-regulatory mechanisms were investigated by comparing Wistar Kyoto rats, which display depression-like behaviors and stress vulnerability, to control Wistar rats. Brain amino acid metabolism in Wistar Kyoto rats was greatly different from normal Wistar rats, with special reference to lower cystathionine and serine levels. In addition, Wistar Kyoto rats demonstrated abnormality in dopamine metabolism compared with Wistar rats. In the case of stress response, amino acid levels having a sedative and/or hypnotic effect were constant in the brain of Wistar Kyoto rats, though these amino acid levels were reduced in Wistar rats under a stressful condition. These results suggest that the abnormal amino acid metabolism may induce depression-like behaviors and stress vulnerability in Wistar Kyoto rats. Therefore, we hypothesized that abnormalities in amino acid and monoamine metabolism may induce depression, and amino acid metabolism in the brain may be related to stress vulnerability.     

Sweet block copolymer nanoparticles: preparation and self-assembly of fully oligosaccharide-based amphiphile

The preparation of biocompatible nanocarriers that have potential applications in the cosmetic and health industries is highly desired. The self-assembly of amphiphilic block copolymers displaying biosourced polysaccharides at the surface is one of the most promising approaches. In the continuity of our works related to the preparation of "hybrid" amphiphilic oligosaccharide-based block copolymers, we present here the design of a new generation of self-assembled nanoparticles composed entirely of oligosaccharide-based amphiphilic block co-oligomers (BCO). These systems are defined by a covalent linkage of the two saccharidic blocks through their reducing end units, resulting in a sweet "head-to-head" connection. As an example, we have prepared and studied a BCO in which the hydrophilic part is composed of a free maltoheptaosyl derivative clicked to a hydrophobic part composed of a peracetylated maltoheptaosyl derivative. This amphiphilic BCO self-assembles to form spherical micelles in water with an average diameter of 30 nm. The efficient enzymatic hydrolysis of the maltoheptaose that constitutes the shell of the micelles was followed by light scattering and colorimetric methods.     

High Risk of ART Non-Adherence and Delay of ART Initiation among HIV Positive Double Orphans in Kigali, Rwanda

Background

To reduce HIV/AIDS related mortality of children, adherence to antiretroviral treatment (ART) is critical in the treatment of HIV positive children. However, little is known about the association between ART adherence and different orphan status. The aims of this study were to assess the ART adherence and identify whether different orphan status was associated with the child’s adherence.

Methods

A total of 717 HIV positive children and the same number of caregivers participated in this cross-sectional study. Children’s adherence rate was measured using a pill count method and those who took 85% or more of the prescribed doses were defined as adherent. To collect data about adherence related factors, we also interviewed caregivers using a structured questionnaire.

Results

Of all children (N = 717), participants from each orphan category (double orphan, maternal orphan, paternal orphan, non-orphan) were 346, 89, 169, and 113, respectively. ART non-adherence rate of each orphan category was 59.3%, 44.9%, 46.7%, and 49.7%, respectively. The multivariate analysis indicated that maternal orphans (AOR 0.31, 95% CI 0.12–0.80), paternal orphans (AOR 0.35, 95% CI 0.14–0.89), and non-orphans (AOR 0.45, 95% CI 0.21–0.99) were less likely to be non-adherent compared to double orphans. Double orphans who had a sibling as a caregiver were more likely to be non-adherent. The first mean CD4 count prior to initiating treatment was 520, 601, 599, and 844 (cells/ml), respectively (p<0.001). Their mean age at sero-status detection was 5.9, 5.3, 4.8, and 3.9 (year old), respectively (p<0.001).

Conclusions

Double orphans were at highest risk of ART non-adherence and especially those who had a sibling as a caregiver had high risk. They were also in danger of initiating ART at an older age and at a later stage of HIV/AIDS compared with other orphan categories. Double orphans need more attention to the promote child’s adherence to ART.     

Heterologous production of bisucaberin using a biosynthetic gene cluster cloned from a deep sea metagenome

A siderophore biosynthetic gene cluster was cloned from a metagenomic library generated from deep sea sediment. The gene cluster was successfully expressed in Escherichia coli to produce bisucaberin, a siderophore originally reported from the marine bacterium Alteromonas haloplanktis. The cloned bisucaberin biosynthetic gene cluster was moderately similar to that of the known bisucaberin producer Vibrio salmonicida. However, the cloned gene cluster consists of four genes rather than three genes found in the V. salmonicida cluster. The low overall homology of the amino acid and nucleotide sequences with those of other species suggests that the cloned genes were derived from one of the unsequenced bacteria including uncultured species.     

The extracellular adenosine deaminase growth factor, ADGF/CECR1, plays a role in Xenopus embryogenesis via the adenosine/P1 receptor

Adenosine deaminase-related growth factors (ADGF), also known as CECR1 in vertebrates, are a novel family of growth factors with sequence similarity to classical cellular adenosine deaminase. Although genes for ADGF/CECR1 have been identified in both invertebrates as well as vertebrates, their in vivo functions in vertebrates remain unknown. We isolated cDNA clones for two cerc 1s from Xenopus laevis. Both recombinant Xenopus CECR1s exhibited adenosine deaminase and growth factor activity, and the adenosine deaminase activity was found to be indispensable for growth factor activity. The Xenopus cerc 1s are expressed in the somites, pronephros, eyes, cement gland, neural tube, and neural floor plate of the embryos. Knock-down of these two genes using morpholino oligonucleotides caused a reduction in the body size and abnormalities of the body axis in the Xenopus embryos, accompanied by selective changes in the expression of developmental marker genes. Injection of adenosine, agonists for adenosine/P1 receptors, or adenosine deaminase inhibitor into late gastrula archenteron embryos resulted in developmental defects similar to those caused by morpholino oligonucleotide injection. These results show, for the first time, the involvement of CECR1s via the adenosine/P1 receptors in vertebrate embryogenesis via regulation of extracellular adenosine concentrations.     

TNF-alpha stimulation inhibits siRNA-mediated RNA interference through a mechanism involving poly-(A) tail stabilization

The control of mRNA stability is a complex biological process that involves numerous factors, including microRNA (miRNA) and short interfering RNA (siRNA). Here, we show that short interfering RNA (siRNA) and microRNA share some similarities in their response to cellular stress. miR16 expedites the degradation of mRNAs containing AU-rich elements (ARE) in their 3' untranslated region (UTR). si20 is an siRNA designed to target a non-ARE sequence in the TNF 3'UTR. We found that both si20 and miR16/ARE-mediated degradation of mRNAs can be inhibited by stimulating cells with different stresses. By analyzing TNF-alpha stimulation-mediated stabilization of si20- and miR16-targeted mRNA, we show that this stabilization is not caused by modifying si20 and miR16 loading into Ago2 complexes, or mRNA targeting to Ago2, but by inhibiting mRNA deadenylation. This is the first report showing that a specific siRNA-mediated mRNA degradation can be regulated by inflammatory stimuli, and that deadenylation is involved in this siRNA-mediated mRNA decay.