A Chloroplast-protein Possessing Photoreducible Disulfide Bond

The five strains of A. fumigatus examined showed marked variation in their ability to produce protease in submerged culture. Also marked differences in protease production were observed when individual strains were tested in different media. Yields of up to 3.7 EU_PH 10.0 per 1 were obtained in a liver-glucose medium with the one pathogenic strain included in the study.     

Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferation of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa.     

Arctic mutant Aβ40 aggregates on α7 nicotinic acetylcholine receptors and inhibits their functions

Amyloid β protein (Aβ) plays a central role in the pathogenesis of Alzheimer's disease (AD). Point mutations within the Aβ sequence associated with familial AD (FAD) are clustered around the central hydrophobic core of Aβ. Several types of mutations within the Aβ sequence have been identified, and the ‘Arctic’ mutation (E22G) has a purely cognitive phenotype typical of AD. Previous studies have shown that the primary result of the ‘Arctic’ mutation is increased formation of Aβ protofibrils. However, the molecular mechanism underlying this effect remains unknown. Aβ42 binds to a neuronal nicotinic acetylcholine receptor subunit, neuronal acetylcholine receptor subunit alpha‐7 (CHRNA7), with high affinity and, thus, may be involved in the pathogenesis of AD. Therefore, to clarify the molecular mechanism of Arctic mutation‐mediated FAD, we focused on CHRNA7 as a target molecule of Arctic Aβ. We performed an in vitro binding assay using purified CHRNA7 and synthetic Arctic Aβ40, and demonstrated that Arctic Aβ40 specifically bound to CHRNA7. The aggregation of Arctic Aβ40 was enhanced with the addition of CHRNA7. Furthermore, the function of CHRNA7 was detected by measuring Ca²⁺ flux and phospho‐p44/42 MAPK (ERK1/2) activation. Our results indicated that Arctic Aβ40 aggregation was enhanced by the addition of CHRNA7, which destabilized the function of CHRNA7 via inhibition of Ca²⁺ responses and activation of ERK1/2. These findings indicate that Arctic Aβ mutation may be involved in the mechanism underlying FAD. This mechanism may involve binding and aggregation, leading to the inhibition of CHRNA7 functions.

     

Cynomolgus monkey cytochrome P450 2C43: cDNA cloning, heterologous expression, purification and characterization

The cDNA of cytochrome P450 (CYP) 2C43 was cloned from cynomolgus monkey liver by RT-PCR. The deduced amino acid sequence showed 93% and 91% identity to human CYP2C9 and CYP2C19, respectively. The cDNA was expressed in Escherichia coli and purified by a series of chromatography steps, yielding a specific content of 11.5 nmol P450/mg protein. The substrate specificity of the purified CYP2C43 was examined in a reconstitution system comprising NADPH-P450 reductase, lipid, cytochrome b(5) and CYP2C marker substrates. The purified CYP2C43 showed high activity for testosterone 17-oxidation and progesterone 21-hydroxylation, which were also observed for CYP2C19 but not CYP2C9. In addition, CYP2C43 showed activity for (S)-mephenytoin 4'-hydroxylation, a marker reaction for CYP2C19. With CYP2C9 marker substrates, CYP2C43 exhibited low activity for diclofenac 4'-hydroxylation and no activity for tolbutamide p-methylhydroxylation. Therefore, in terms of substrate specificity, our results indicate that CYP2C43 is similar to CYP2C19, rather than CYP2C9.     

Plasmodium falciparum: Development and validation of a measure of intraerythrocytic growth using SYBR Green I in a flow cytometer

Reliable analytical techniques to test growth-promoting and antimalarial efficacy on plasmodia are very important. Flow cytometry (FCM) offers the possibility to study developmental stages of intraerythrocytic growth of malaria parasites using nucleic acid staining. To analyze the growth of Plasmodium falciparum SYBR Green I was introduced as an intercalating dye with FCM for the 488 nm line of an argon laser. Procedures employing FCM, including fixatives, dye concentrations, dilution buffer, and staining period, were optimized to simplify the method. FCM as described here allows parasitemia and parasites of different stages to be quantified according to the DNA content. The proportion of parasitized erythrocytes estimated by FCM and the Giemsa method agreed with determination by parasite lactate dehydrogenase. The protocol was extended to merozoite counting as a sensitive assay of growth inhibition of the parasite.     

Efficacy of genotype notification to Japanese smokers on smoking cessation—An intervention study at workplace

Objectives: It is well-known that smoking causes many diseases including cancers. Informing smokers of their genotypes associated with the vulnerability to the harms of smoking may be effective measures for smoking cessation. The present study examined the effects of genotype notification of an oncogene (L-myc) genotype to smokers on their behavior to quit smoking. Methods: Subjects were 562 employees of a bank who answered to be a smoker for a questionnaire used at annual health checkup at workplace from July to December 2002. Those enrolled on August, October, and December were allocated into the genotype notification group (intervention group), and the rest into the controls. Among 286 smokers allocated into the intervention group, 257 participants (89.9%) agreed to genotype testing. One year after the enrollment, a follow-up questionnaire survey was conducted for all smokers including controls. Results: Those who stated to have quitted smoking were 22 (8.0%) among the 276 controls and 15 (5.8%) among the 257 genotype notified participants, providing that the odds ratio (OR) of cessation for the intervention was 0.64 (95% confidence interval, 0.32–1.28). No psychological problems associated with genotype notification were observed. Conclusion: The present study did not show positive effects of genotype notification on smoking cessation rate. To elevate the cessation rate, methods to explain and notify genotypes should be improved.     

Preparation, characterization, and biological evaluation of 6(I),6(IV)-di-O-[α-l-fucopyranosyl-(1→6)-2-acetamido-2-deoxy-β-d-glucopyranosyl]-cyclomaltoheptaose and 6-O-[α-l-fucopyranosyl-(1→6)-2-acetamido-2-deoxy-β-d-glucopyranosyl]-cyclomaltoheptaose

6(I),6(IV)-Di-O-[α-l-fucopyranosyl-(1→6)-2-acetamido-2-deoxy-β-d-glucopyranosyl]-cyclomaltoheptaose (βCD) {6(I),6(IV)-di-O-[α-l-Fuc-(1→6)-β-d-GlcNAc]-βCD (5)} and 6-O-[α-l-fucopyranosyl-(1→6)-2-acetamido-2-deoxy-β-d-glucopyranosyl]-βCD {6-O-[α-l-Fuc-(1→6)-β-d-GlcNAc]-βCD (6)} were chemically synthesized using the corresponding authentic compounds, bis(2,3-di-O-acetyl)-pentakis(2,3,6-tri-O-acetyl)-βCD as the glycosyl acceptor and 2,3,4-tri-O-benzyl-α-l-fucopyranosyl-(1→6)-3,4-di-O-acetyl-2-deoxy-2-(2,2,2-trichloroethoxycarbonylamino)-d-glucopyranosyl trichloroacetimidate as the fuco-glucosaminyl donor. NMR confirmed that α-l-Fuc-(1→6)-d-GlcNAc was bonded by β-linking to the βCD ring. To evaluate biological efficiency, the biological activities of the new branched βCDs were examined. The cell detachment activity of 5 was lower than that of 6 in real-time cell sensing (RT-CES) assay, indicating that 5 has lower toxicity. In SPR analysis, 5 had a higher special binding with AAL, a fucose-recognizing lectin. These results suggest that 5 could be an efficient drug carrier directed at cells expressing fucose-binding proteins.     

Association between Combined Lifestyle Factors and Non-Restorative Sleep in Japan: A Cross-Sectional Study Based on a Japanese Health Database

Background

Although lifestyle factors such as cigarette smoking, excessive drinking, obesity, low or no exercise, and unhealthy dietary habits have each been associated with inadequate sleep, little is known about their combined effect. The aim of this study was to quantify the overall impact of lifestyle-related factors on non-restorative sleep in the general Japanese population.

Methods and Findings

A cross-sectional study of 243,767 participants (men, 39.8%) was performed using the Specific Health Check and Guidance System in Japan. A healthy lifestyle score was calculated by adding up the number of low-risk lifestyle factors for each participant. Low risk was defined as (1) not smoking, (2) body mass index<25 kg/m², (3) moderate or less alcohol consumption, (4) regular exercise, and (5) better eating patterns. Logistic regression analysis was used to examine the relationship between the score and the prevalence of non-restorative sleep, which was determined from questionnaire responses. Among 97,062 men (mean age, 63.9 years) and 146,705 women (mean age, 63.7 years), 18,678 (19.2%) and 38,539 (26.3%) reported non-restorative sleep, respectively. The prevalence of non-restorative sleep decreased with age for both sexes. Compared to participants with a healthy lifestyle score of 5 (most healthy), those with a score of 0 (least healthy) had a higher prevalence of non-restorative sleep (odds ratio, 1.59 [95% confidence interval, 1.29–1.97] for men and 2.88 [1.74–4.76] for women), independently of hypertension, hypercholesterolemia, diabetes, and chronic kidney disease. The main limitation of the study was the cross-sectional design, which limited causal inferences for the identified associations.

Conclusions

A combination of several unhealthy lifestyle factors was associated with non-restorative sleep among the general Japanese population. Further studies are needed to establish whether general lifestyle modification improves restorative sleep.     

A new class of type I protein geranylgeranyltransferase (GGTase I) inhibitor

Replacement of the thiol groups in 1, a potent and highly selective Candida albicans GGTase I inhibitor discovered through screening, with an imidazole ring was achieved by using solid phase synthesis. A non-thiol compound, 7, was found as a representative of a new class of potent C. albicans GGTase I inhibitor with high selectivity against human GGTase I.     

Dissociable anterograde amnesic effects of retrosplenial cortex and hippocampal lesions on spontaneous object recognition memory in rats

The amnesic effects of excitotoxic lesions of the rat retrosplenial cortex (RS) and hippocampus (HPC) in the spontaneous object recognition (SOR) performance were investigated. The SOR test consisted of the sample-exposure session(s) and a test session. First, to test retrograde amnesia, rats received four sample-exposure sessions within a day at 4 weeks and 1 day before the surgery, respectively. In the test sessions conducted 1 week after the surgery, both lesion groups showed a temporally ungraded retrograde amnesia. Second, to test anterograde amnesia, 1- and 4-week retention intervals were inserted between the four sample-exposure sessions and the test session. The RS-lesioned rats showed a retention interval-dependent impairment in the test sessions, while the HPC-lesioned rats showed an impairment regardless of the retention interval. Finally, to test short-term recognition memory, 5- or 30-min delay was interposed between the single sample-exposure session and the test session. Both lesion groups performed normally irrespective of the delay length. These results suggest that both the RS and HPC are important for long-term object recognition memory, but these areas have different roles in it. ? 2012 Wiley Periodicals, Inc.