Association between delayed bedtime and sleep-related problems among community-dwelling 2-year-old children in Japan

Background

Although delayed sleep timing causes many socio-psycho-biological problems such as sleep loss, excessive daytime sleepiness, obesity, and impaired daytime neurocognitive performance in adults, there are insufficient data showing the clinical significance of a ‘night owl lifestyle’ in early life. This study examined the association between habitual delayed bedtime and sleep-related problems among community-dwelling 2-year-old children in Japan.

Methods

Parents/caregivers of 708 community-dwelling 2-year-old children in Nishitokyo City, Tokyo, participated in the study. The participants answered a questionnaire to evaluate their child’s sleep habits and sleep-related problems for the past 1 month.

Results

Of the 425 children for whom complete data were collected, 90 (21.2%) went to bed at 22:00 or later. Children with delayed bedtime showed significantly more irregular bedtime, delayed wake time, shorter total sleep time, and difficulty in initiating and terminating sleep. Although this relationship indicated the presence of sleep debt in children with delayed bedtime, sleep onset latency did not differ between children with earlier bedtime and those with delayed bedtime. Rather, delayed bedtime was significantly associated with bedtime resistance and problems in the morning even when adjusting for nighttime and daytime sleep time.

Conclusions

Even in 2-year-old children, delayed bedtime was associated with various sleep-related problems. The causal factors may include diminished homeostatic sleep drive due to prolonged daytime nap as well as diurnal preference (morning or night type) regulated by the biological clock.     

Design and Evaluation of the Highly Concentrated Human IgG Formulation Using Cyclodextrin Polypseudorotaxane Hydrogels

To achieve the potent therapeutic effects of human immunoglobulin G (IgG), highly concentrated formulations are required. However, the stabilization for highly concentrated human IgG is laborious work. In the present study, to investigate the potentials of polypseudorotaxane (PPRX) hydrogels consisting of polyethylene glycol (PEG) and α- or γ-cyclodextrin (α- or γ-CyD) as pharmaceutical materials for highly concentrated human IgG, we designed the PPRX hydrogels including human IgG and evaluated their pharmaceutical properties. The α- and γ-CyDs formed PPRX hydrogels with PEG (M.W. 20,000) even in the presence of highly concentrated human IgG (>100 mg/mL). According to the results of ¹H-NMR, powder X-ray diffraction, and Raman microscopy, the formation of human IgG/CyD PPRX hydrogels was based on physical cross-linking arising from their columnar structures. The release profiles of human IgG from the hydrogels were in accordance with the non-Fickian diffusion model. Importantly, the stabilities of human IgG included into the hydrogels against thermal and shaking stresses were markedly improved. These findings suggest that PEG/CyD PPRX hydrogels are useful to prepare the formulation for highly concentrated human IgG.KEY WORDS: cyclodextrin, human IgG, hydrogel, polypseudorotaxane, stability     

Epigenetic regulation of autophagy by the methyltransferase EZH2 through an MTOR-dependent pathway

Macroautophagy is an evolutionarily conserved cellular process involved in the clearance of proteins and organelles. Although the autophagy regulation machinery has been widely studied, the key epigenetic control of autophagy process still remains unknown. Here we report that the methyltransferase EZH2 (enhancer of zeste 2 polycomb repressive complex 2 subunit) epigenetically represses several negative regulators of the MTOR (mechanistic target of rapamycin [serine/threonine kinase]) pathway, such as TSC2, RHOA, DEPTOR, FKBP11, RGS16 and GPI. EZH2 was recruited to these genes promoters via MTA2 (metastasis associated 1 family, member 2), a component of the nucleosome remodeling and histone deacetylase (NuRD) complex. MTA2 was identified as a new chromatin binding protein whose association with chromatin facilitated the subsequent recruitment of EZH2 to silenced targeted genes, especially TSC2. Downregulation of TSC2 (tuberous sclerosis 2) by EZH2 elicited MTOR activation, which in turn modulated subsequent MTOR pathway-related events, including inhibition of autophagy. In human colorectal carcinoma (CRC) tissues, the expression of MTA2 and EZH2 correlated negatively with expression of TSC2, which reveals a novel link among epigenetic regulation, the MTOR pathway, autophagy induction, and tumorigenesis.     

Effectiveness of Trivalent Inactivated Influenza Vaccine in Children Estimated by a Test-Negative Case-Control Design Study Based on Influenza Rapid Diagnostic Test Results

We assessed vaccine effectiveness (VE) against medically attended, laboratory-confirmed influenza in children 6 months to 15 years of age in 22 hospitals in Japan during the 2013–14 season. Our study was conducted according to a test-negative case-control design based on influenza rapid diagnostic test (IRDT) results. Outpatients who came to our clinics with a fever of 38°C or over and had undergone an IRDT were enrolled in this study. Patients with positive IRDT results were recorded as cases, and patients with negative results were recorded as controls. Between November 2013 and March 2014, a total of 4727 pediatric patients (6 months to 15 years of age) were enrolled: 876 were positive for influenza A, 66 for A(H1N1)pdm09 and in the other 810 the subtype was unknown; 1405 were positive for influenza B; and 2445 were negative for influenza. Overall VE was 46% (95% confidence interval [CI], 39–52). Adjusted VE against influenza A, influenza A(H1N1)pdm09, and influenza B was 63% (95% CI, 56–69), 77% (95% CI, 59–87), and 26% (95% CI, 14–36), respectively. Influenza vaccine was not effective against either influenza A or influenza B in infants 6 to 11 months of age. Two doses of influenza vaccine provided better protection against influenza A infection than a single dose did. VE against hospitalization influenza A infection was 76%. Influenza vaccine was effective against influenza A, especially against influenza A(H1N1)pdm09, but was much less effective against influenza B.     

Comparative Phylogenetic Studies on Schistosoma japonicum and Its Snail Intermediate Host Oncomelania hupensis: Origins,Dispersal and Coevolution

Background

Schistosoma japonicum causes major public health problems in China and the Philippines; this parasite, which is transmitted by freshwater snails of the species Oncomelania hupensis, causes the disease intestinal schistosomiasis in humans and cattle. Researchers working on Schistosoma in Africa have described the relationship between the parasites and their snail intermediate hosts as coevolved or even as an evolutionary arms race. In the present study this hypothesis of coevolution is evaluated for S. japonicum and O. hupensis. The origins and radiation of the snails and the parasite across China, and the taxonomic validity of the sub-species of O. hupensis, are also assessed.

Methodology/Principal Findings

The findings provide no evidence for coevolution between S. japonicum and O. hupensis, and the phylogeographical analysis suggests a heterochronous radiation of the parasites and snails in response to different palaeogeographical and climatic triggers. The results are consistent with a hypothesis of East to West colonisation of China by Oncomelania with a re-invasion of Japan by O. hupensis from China. The Taiwan population of S. japonicum appears to be recently established in comparison with mainland Chinese populations.

Conclusions/Significance

The snail and parasite populations of the western mountain region of China (Yunnan and Sichuan) appear to have been isolated from Southeast Asian populations since the Pleistocene; this has implications for road and rail links being constructed in the region, which will breach biogeographical barriers between China and Southeast Asia. The results also have implications for the spread of S. japonicum. In the absence of coevolution, the parasite may more readily colonise new snail populations to which it is not locally adapted, or even new intermediate host species; this can facilitate its dispersal into new areas. Additional work is required to assess further the risk of spread of S. japonicum.     

Knockdown of aquaporin-8 induces mitochondrial dysfunction in 3T3-L1 cells

BackgroundAquaporin-8 (AQP8), a member of the aquaporin water channel family, is expressed in various tissue and cells, including liver, testis, and pancreas. AQP8 appears to have functions on the plasma membrane and/or on the mitochondrial inner membrane. Mitochondrial AQP8 with permeability for water, H₂O₂ and NH₃ has been expected to have important role in various cells, but its information is limited to a few tissues and cells including liver and kidney. In the present study, we found that AQP8 was expressed in the mitochondria in mouse adipose tissues and 3T3-L1 preadipocytes, and investigated its role by suppressing its gene expression.MethodsAQP8-knocked down (shAQP8) cells were established using a vector expressing short hairpin RNA. Cellular localization of AQP8 was examined by western blotting and immunocytochemistry. Mitochondrial function was assessed by measuring mitochondrial membrane potential, oxygen consumption and ATP level measurements.ResultsIn 3T3-L1 cells, AQP8 was expressed in the mitochondria. In shAQP8 cells, mRNA and protein levels of AQP8 were decreased by about 75%. The shAQP8 showed reduced activities of complex IV and ATP synthase; it is probable that the impaired mitochondrial water handling in shAQP8 caused suppression of the electron transport and ADP phosphorylation through inhibition of the two steps which yield water. The reduced activities of the last two steps of oxidative phosphorylation in shAQP8 cause low routine and maximum capacity of respiration and mitochondrial hyperpolarization.ConclusionMitochondrial AQP8 contributes to mitochondrial respiratory function probably through maintenance of water homeostasis.General significanceThe AQP8-knocked down cells we established provides a model system for the studies on the relationships between water homeostasis and mitochondrial function.     

Functional Characterization of the Vitamin K2 Biosynthetic Enzyme UBIAD1

UbiA prenyltransferase domain-containing protein 1 (UBIAD1) plays a significant role in vitamin K₂ (MK-4) synthesis. We investigated the enzymological properties of UBIAD1 using microsomal fractions from Sf9 cells expressing UBIAD1 by analysing MK-4 biosynthetic activity. With regard to UBIAD1 enzyme reaction conditions, highest MK-4 synthetic activity was demonstrated under basic conditions at a pH between 8.5 and 9.0, with a DTT ≥0.1 mM. In addition, we found that geranyl pyrophosphate and farnesyl pyrophosphate were also recognized as a side-chain source and served as a substrate for prenylation. Furthermore, lipophilic statins were found to directly inhibit the enzymatic activity of UBIAD1. We analysed the aminoacid sequences homologies across the menA and UbiA families to identify conserved structural features of UBIAD1 proteins and focused on four highly conserved domains. We prepared protein mutants deficient in the four conserved domains to evaluate enzyme activity. Because no enzyme activity was detected in the mutants deficient in the UBIAD1 conserved domains, these four domains were considered to play an essential role in enzymatic activity. We also measured enzyme activities using point mutants of the highly conserved aminoacids in these domains to elucidate their respective functions. We found that the conserved domain I is a substrate recognition site that undergoes a structural change after substrate binding. The conserved domain II is a redox domain site containing a CxxC motif. The conserved domain III is a hinge region important as a catalytic site for the UBIAD1 enzyme. The conserved domain IV is a binding site for Mg²⁺/isoprenyl side-chain. In this study, we provide a molecular mapping of the enzymological properties of UBIAD1.     

Precocene II,a Trichothecene Production Inhibitor,Binds to Voltage-Dependent Anion Channel and Increases the Superoxide Level in Mitochondria of Fusarium graminearum

Precocene II, a constituent of essential oils, shows antijuvenile hormone activity in insects and inhibits trichothecene production in fungi. We investigated the molecular mechanism by which precocene II inhibits trichothecene production in Fusarium graminearum, the main causal agent of Fusarium head blight and trichothecene contamination in grains. Voltage-dependent anion channel (VDAC), a mitochondrial outer membrane protein, was identified as the precocene II-binding protein by an affinity magnetic bead method. Precocene II increased the superoxide level in mitochondria as well as the amount of oxidized mitochondrial proteins. Ascorbic acid, glutathione, and α-tocopherol promoted trichothecene production by the fungus. These antioxidants compensated for the inhibitory activity of precocene II on trichothecene production. These results suggest that the binding of precocene II to VDAC may cause high superoxide levels in mitochondria, which leads to stopping of trichothecene production.     

IL-27 affects helper T cell responses via regulation of PGE2 production by macrophages

IL-27 is a heterodimeric cytokine that regulates both innate and adaptive immunity. The immunosuppressive effect of IL-27 largely depends on induction of IL-10-producing Tr1 cells. To date, however, effects of IL-27 on regulation of immune responses via mediators other than cytokines remain poorly understood. To address this issue, we examined immunoregulatory effects of conditional medium of bone marrow-derived macrophages (BMDMs) from WSX-1 (IL-27Rα)-deficient mice and found enhanced IFN-γ and IL-17A secretion by CD4⁺ T cells as compared with that of control BMDMs. We then found that PGE₂ production and COX-2 expression by BMDMs from WSX-1-deficient mice was increased compared to control macrophages in response to LPS. The enhanced production of IFN-γ and IL-17A was abolished by EP2 and EP4 antagonists, demonstrating PGE₂ was responsible for enhanced cytokine production. Murine WSX-1-expressing Raw264.7 cells (mWSX-1-Raw264.7) showed phosphorylation of both STAT1 and STAT3 in response to IL-27 and produced less amounts of PGE₂ and COX-2 compared to parental RAW264.7 cells. STAT1 knockdown in parental RAW264.7 cells and STAT1-deficiency in BMDMs showed higher COX-2 expression than their respective control cells. Collectively, our result indicated that IL-27/WSX-1 regulated PGE₂ secretion via STAT1–COX-2 pathway in macrophages and affected helper T cell response in a PGE₂-mediated fashion.