A MicroRNA Derived from Adenovirus Virus-Associated RNAII Promotes Virus Infection via Posttranscriptional Gene Silencing.

The adenovirus (Ad) serotype 5 genome encodes two noncoding small RNAs (virus-associated RNAs I and II [VA-RNAI and -II]), which are approximately 160-nucleotide (nt) RNAs transcribed by RNA polymerase III. It is well known that VA-RNAI supports Ad infection via the inhibition of double-stranded RNA-dependent protein kinase (PKR), which recognizes double-stranded RNA and acts as an antiviral system. Recent studies revealed that VA-RNAs are processed into VA-RNA-derived microRNAs (miRNAs) (mivaRNAI and -II); however, we and another group recently demonstrated that mivaRNAI does not promote Ad replication. On the other hand, the roles of VA-RNAII and mivaRNAII in Ad replication have remained to be clarified. In this study, we demonstrated mivaRNAII-mediated promotion of Ad replication. Transfection with chemically synthesized 3'-mivaRNAII-138, one of the most abundant forms of mivaRNAII, significantly enhanced Ad replication, while the other species of mivaRNAII did not. We identified 8 putative target genes of 3'-mivaRNAII-138 by microarray analysis and in silico analysis. Among the 8 candidates, knockdown of the cullin 4A (CUL4A) gene, which encodes a component of the ubiquitin ligase complex, most significantly enhanced Ad replication. CUL4A expression was significantly suppressed by 3'-mivaRNAII-138 via posttranscriptional gene silencing, indicating that CUL4A is a target gene of 3'-mivaRNAII-138 and mivaRNAII functions as a viral miRNA promoting Ad infection. It has been reported that CUL4A is involved in degradation of c-Jun, which acts as a transcription factor in the Jun-N-terminal kinase (JNK) signaling cascade. Treatment with JNK inhibitors dramatically suppressed Ad replication, suggesting that mivaRNAII-mediated downregulation of CUL4A enhanced JNK signaling and thereby promoted Ad infection.IMPORTANCE Several types of viruses encode viral miRNAs which regulate host and/or viral gene expression via posttranscriptional gene silencing, leading to efficient viral infection. Adenovirus (Ad) expresses miRNAs derived from VA-RNAs (mivaRNAI and -II); however, recent studies have revealed that processing of VA-RNAI into mivaRNAI inhibits Ad replication. Conversely, we demonstrate here that mivaRNAII significantly promotes Ad replication and that mivaRNAII-mediated suppression of CUL4A expression via posttranscriptional gene silencing induces accumulation of c-Jun, leading to promotion of Ad infection. These results exhibited the significance of VA-RNAII for supporting Ad infection through a mechanism complementary to that of VA-RNAI. These observations could provide important clues toward a new perspective on host-virus interaction. Moreover, Ad is widely used as a basic framework for viral vectors and oncolytic viruses. Our findings will help to regulate Ad infection and will promote the development of novel Ad vectors and oncolytic Ad.

Axillary lymph-node metabolic activity assessment on 18F-FDG-PET/CT in rheumatoid arthritis patients treated with biologic therapies.

Objective: Recent studies have provided new insights into the role of lymph nodes (LNs) in rheumatoid arthritis (RA). The aim of this study was to evaluate the metabolic activity of the axillary LNs in relation to that of the upper limb joints and the clinical assessment of disease activity in RA patients treated with biologic therapies.Method: 18F-fluorodeoxyglucose-positron emission tomography/computed tomography (18F-FDG-PET/CT) scans were acquired for 64 patients with RA at baseline and after 6 months of biologic therapy, and the patients' clinical status was evaluated. The maximum standardized uptake value (SUVmax), metabolic active volume, and total lesion glycolysis (TLG) were used to assess glucose metabolism in the LNs and 12 joints. Clinical evaluations included serum markers and the Disease Activity Score based on 28-joint count-erythrocyte sedimentation rate (DAS28-ESR).Results: Changes in the SUVmax and TLG for the axillary LNs correlated significantly with those of the ipsilateral wrist joints. There was a positive correlation between the changes in the three metabolic parameters of the axillary LNs and the changes in disease activity after treatment. After 6 months of biologic therapy, all metabolic parameters for the axillary LNs in patients with a DAS28-ESR < 3.2 were significantly lower than those of patients with a DAS28-ESR ≥ 3.2.Conclusion: A relationship between the glucose metabolism of the axillary LNs and the ipsilateral wrist joints was demonstrated by the 18F-FDG-PET/CT parameters. The metabolic activity and active volume of axillary LNs may reflect the therapeutic response to the biologic treatment of RA.

Phase Competition in the Palmer-Chalker XY Pyrochlore Er_{2}Pt_{2}O_{7}.

We report neutron scattering measurements on Er_{2}Pt_{2}O_{7}, a new addition to the XY family of frustrated pyrochlore magnets. Symmetry analysis of our elastic scattering data shows that Er_{2}Pt_{2}O_{7} orders into the k=0, Γ_{7} magnetic structure (the Palmer-Chalker state), at T_{N}=0.38 K. This contrasts with its sister XY pyrochlore antiferromagnets Er_{2}Ti_{2}O_{7} and Er_{2}Ge_{2}O_{7}, both of which order into Γ_{5} magnetic structures at much higher temperatures, T_{N}=1.2 and 1.4 K, respectively. In this temperature range, the magnetic heat capacity of Er_{2}Pt_{2}O_{7} contains a broad anomaly centered at T^{*}=1.5 K. Our inelastic neutron scattering measurements reveal that this broad heat capacity anomaly sets the temperature scale for strong short-range spin fluctuations. Below T_{N}=0.38 K, Er_{2}Pt_{2}O_{7} displays a gapped spin-wave spectrum with an intense, flat band of excitations at lower energy and a weak, diffusive band of excitations at higher energy. The flat band is well described by classical spin-wave calculations, but these calculations also predict sharp dispersive branches at higher energy, a striking discrepancy with the experimental data. This, in concert with the strong suppression of T_{N}, is attributable to enhanced quantum fluctuations due to phase competition between the Γ_{7} and Γ_{5} states that border each other within a classically predicted phase diagram.

Associations of sleep bruxism with age, sleep apnea, and daytime problematic behaviors in children.

OBJECTIVES: The aims of this study were to investigate the prevalence of sleep bruxism in children in Japan, and its relationships with sleep-related factors and daytime problematic behavior. SUBJECTS AND METHODS: Guardians of 6023 children aged 2-12 years completed the Japanese Sleep Questionnaire. Multiple regression analysis and structural equation modeling were performed. RESULTS: Sleep bruxism was reported in 21.0% children (n = 1263): the prevalence was highest in the age group of 5-7 years (27.4%). Multiple regression analysis showed that sleep bruxism had significant correlations with age 5-7 years (OR: 1.72; P < 0.0001), 'Moves a lot during sleep' (OR: 1.47; P < 0.0001), 'sleeps with mouth open' (OR: 1.56; P < 0.0001), and 'snores loudly' (OR: 1.80; P < 0.0001). In structural equation modeling, sleep bruxism had a significant but weak direct effect on daytime problematic behavior, while sleep bruxism significantly correlated with obstructive sleep apnea, which had a higher direct effect on daytime problematic behavior. CONCLUSIONS: Sleep bruxism was reported in 21.0% of Japanese children and had independent relationships with age, movements during sleep, and snoring. A comorbidity of sleep-disordered breathing might be related to daytime problematic behavior in children with sleep bruxism.

Sex and the clinical value of body mass index in patients with clear cell renal cell carcinoma.

BACKGROUND: An increased body mass index (BMI) is significantly associated with favourable prognosis in renal cell carcinoma (RCC). This study investigated the associations among sex, BMI, and prognosis in clear cell RCC patients. METHODS: We retrospectively analysed 435 patients with clear cell RCC who underwent a nephrectomy. The associations among sex, BMI, clinicopathologic factors, and cancer-specific survival (CSS) were analysed. RESULTS: As a continuous variable, increased BMI was associated with higher CSS rate by univariate analysis in the whole population (hazard ratio, 0.888 per kg m(-2); 95% confidence interval, 0.803-0.982; P=0.021). A sub-population analysis by sex demonstrated that BMI was significantly associated with CSS in men (P=0.004) but not in women (P=0.725). Multivariate analysis revealed BMI to be an independent predictor of CSS in only men. CONCLUSION: Body mass index was significantly associated with clear cell RCC prognosis. However, the clinical value of BMI may be different between men and women.

Epistatic relationship between Waardenburg syndrome genes MITF and PAX3.

Waardenburg syndrome (WS) is a hereditary disorder that causes hypopigmentation and hearing impairment. Depending on additional symptoms, WS is classified into four types: WS1, WS2, WS3 and WS4. Mutations in MITF (microphthalmia-associated transcription factor) and PAX3, encoding transcription factors, are responsible for WS2 and WS1/WS3, respectively. We have previously shown that MITF transactivates the gene for tyrosinase, a key enzyme for melanogenesis, and is critically involved in melanocyte differentiation. Absence of melanocytes affects pigmentation in the skin, hair and eyes, and hearing function in the cochlea. Therefore, hypopigmentation and hearing loss in WS2 are likely to be the results of an anomaly of melanocyte differentiation caused by MITF mutations. However, the molecular mechanism by which PAX3 mutations cause the auditory-pigmentary symptoms in WS1/WS3 remains to be explained. Here we show that PAX3, a transcription factor with a paired domain and a homeodomain, transactivates the MITF promoter. We further show that PAX3 proteins associated with WS1 in either the paired domain or the homeodomain fail to recognize and transactivate the MITF promoter. These results provide evidence that PAX3 directly regulates MITF and suggest that the failure of this regulation due to PAX3 mutations causes the auditory-pigmentary symptoms in at least some individuals with WS1.

Ectopic expression of MITF, a gene for Waardenburg syndrome type 2, converts fibroblasts to cells with melanocyte characteristics.

MITF (microphthalmia-associated transcription factor) encodes a transcription factor with a basic-helix-loop-helix-zipper (bHLH-Zip) motif. MITF mutations occur in patients with Waardenburg syndrome type 2, a disorder associated with melanocyte abnormalities. Here we show that ectopic expression of MITF converts NIH/3T3 fibroblasts into cells with characteristics of melanocytes. MITF transfectants formed foci of morphologically altered cells, which resemble those induced by oncogenes, but did not exhibit malignant phenotypes. Instead, they contained dendritic cells that express melanogenic marker proteins such as tyrosinase and tyrosinase-related protein 1. Most cloned cells of MITF transfectants exhibited dendritic morphology and expressed melanogenic markers, but such properties were not observed in cells transfected with closely related TFE3 cDNA. Our findings indicate that MITF is critically involved in melanocyte differentiation.

Persistence of ferroelectricity in BaTiO3 through the insulator-metal transition.

The ferroelectric BaTiO(3) is a band-gap insulator. Itinerant electrons can be introduced in this material by doping, for example, with oxygen vacancies. Above a critical electron concentration of n(c) approximately 1 x 10(20) cm(-3), BaTiO(3-delta) becomes metallic. This immediately raises a question: Does metallic BaTiO(3-delta) still retain ferroelectricity? One may expect itinerant electrons to destroy ferroelectricity as they screen the long-range Coulomb interactions. We followed the phase transitions in BaTiO(3-delta) as a function of n far into metallic phase. Although their stability range decreases with n, the low-symmetry phases in metallic BaTiO(3-delta) are still retained up to an estimated concentration of n* approximately 1.9 x 10(21) cm(-3). Moreover, it appears that the itinerant electrons partially stabilize the ferroelectric phases in metallic BaTiO(3-delta) by screening strong crystal field perturbations caused by oxygen vacancies.

Annexin II represents metastatic potential in clear-cell renal cell carcinoma.

BACKGROUND: Annexin II (ANX2) is a multi-functional protein involved in cell proliferation and membrane physiology and is related to cancer progression. The purpose of this study was to assess ANX2 expression in clear-cell (cc) renal cell carcinoma (RCC). METHODS: The ANX2 expression in 18 primary ccRCCs was examined by real-time reverse transcriptase (RT)-PCR and western blot analyses. Furthermore, immunohistochemical study was performed using paraffin section of 154 primary ccRCCs and 24 metastases. The association between ANX2 expression and the clinicopathological factors and prognosis was analysed. RESULTS: The ANX2 was upregulated at both mRNA and protein levels in 14 of 18 primary ccRCCs. Immunohistochemical analysis showed that ANX2 was positive in 73 (47.4%) of 154 primary ccRCC and in 21 (87.5%) of 24 metastatic tumours. The ANX2 expression in the primary tumours showed significant associations with a higher stage, a higher nuclear grade. In patients without metastasis, the 5-year metastasis-free rate in patients with ANX2-positive tumour was significantly lower than that in those with ANX2-negative tumour (63.0% vs 90.1%; P<0.0001). Multivariate analysis showed that ANX2 expression is an independent predictor for metastasis. CONCLUSION: Our findings suggest that ANX2 expression might be a novel predictor of the metastatic potential of ccRCC.

Neuronal architecture of on and off pathways to ganglion cells in carp retina.

Bipolar, amacrine, and ganglion cells of carp retina, stained intracellularly with Procion yellow, can be divided into types a and b, according to the destination of terminals and dendritic trees in the inner plexiform layer (sublamina a and b, respectively). Type a cells showed hyperpolarizing, or off, responses and type b cells depolarizing, or on, responses. Carp thus resembles cat in the basic organization of on and off pathways in the retina.

Detection of an auditory nerve--activating substance.

A substance or substances capable of increasing the firing rate of primary auditory fibers is detectable in the perilymph of frogs and guinea pigs subjected to sound stimulation. The increase in firing rate occurs in single units of the frog auditory nerve after perilymph obtained from frogs or guinea pigs during sound stimulation is infused into the frog perilymphatic sac. Perilymph collected from animals maintained in silence failed to cause an increase in firing rate of primary auditory fibers of the frog.

Elastic constants in tetragonal hen egg-white lysozyme crystals containing large amount of water.

Transverse sound velocity of cross-linked tetragonal hen egg-white (HEW) lysozyme crystals containing large amount of water in the crystal was measured using ultrasonic pulse-echo method. All elastic constants of cross-linked crystals were observed to be C11=C22=5.50 GPa, C12=4.33 GPa, C13=C23=3.94 GPa, C33=5.22 GPa, C44=C55=0.68 GPa, and C66=0.84 GPa, respectively. We found that the elastic constants of the cross-linked crystals are identical to those of the intrinsic ones without cross-linking. Moreover, we found that tetragonal HEW lysozyme crystals that enclose large amount of water show decreased elastic constants (softening). In particular, the shear elastic constants C44=C55 and C66 showed more softening effect comparing with other elastic components.

Ectopic (subcutaneous) Paragonimus miyazakii infection in a dog.

Ectopic infection with Paragonimus miyazakii was determined to be the cause of a subcutaneous inguinal mass in a 15-month-old, male, boar-hunting dog. On histologic examination, the mass comprised granulomatous panniculitis, intralesional adult trematodes and eggs, and lymphadenitis. Extrapulmonary paragonimosis in animals is rare. This appears to be the first report in a dog of ectopic P. miyazakii infection with mature trematodes and eggs that involved the inguinofemoral lymphocenter and surrounding subcutis.

Oral medicine psychiatric liaison clinic: study of 1202 patients attending over an 18-year period.

Patients with orofacial pain and discomfort often suffer from psychiatric disorders. However, few studies involving a large sample have examined the diagnostic results of patients with orofacial pain or discomfort in relation to psychiatric disorders. The purpose of this study was to summarize and clarify the characteristics and demographic data of 1202 patients attending the psychiatric liaison clinic at Aichi Gakuin University Hospital. Psychiatric diagnosis was performed by psychiatrists for all patients, based on the Diagnostic and Statistical Manual of Mental Disorders, fifth edition. Among the 1202 patients, 992 (82.5%) were female. The average age of the patients was 57.2±15.0years. The predominant broad categories of orofacial pain and discomfort seen were burning mouth syndrome (n=484, 40.3%), persistent idiopathic facial pain (n=258, 21.5%), and oral dysesthesia (n=215, 17.9%). The predominant broad categories of psychiatric diagnoses seen were somatic symptoms and related disorders (n=934, 77.7%) and depressive disorders (n=76, 6.3%). Among the 934 patients with somatic symptoms and related disorders, 678 had a somatic symptom disorder with predominant pain. The results confirmed that most patients with orofacial pain and discomfort were middle-aged and elderly women suffering from a somatic symptom disorder with predominant pain.

Submillisecond kinetics of glutamate release from a sensory synapse.

Exocytosis-mediated glutamate release from ribbon-type synaptic terminals of retinal bipolar cells was studied using AMPA receptors and simultaneous membrane capacitance measurements. Release onset (delay <0.8 ms) and offset were closely tied to Ca2+ channel opening and closing. Asynchronous release was not copious and we estimate that there are approximately 5 Ca2+ channels per docked synaptic vesicle. Depending on Ca2+ current amplitude, release occurred in a single fast bout or in two successive bouts with fast and slow onset kinetics. The second, slower bout may reflect a mobilization rate of reserve vesicles toward fusion sites that is accelerated by increasing Ca2+ influx. Bipolar cell synaptic ribbons thus are remarkably versatile signal transducers, capable of transmitting rapidly changing sensory input, as well as sustained stimuli, due to their large pool of releasable vesicles.

A key role of starburst amacrine cells in originating retinal directional selectivity and optokinetic eye movement.

The directional selectivity of retinal ganglion cell responses represents a primitive pattern recognition that operates within a retinal neural circuit. The cellular origin and mechanism of directional selectivity were investigated by selectively eliminating retinal starburst amacrine cells, using immunotoxin-mediated cell targeting techniques. Starburst cell ablation in the adult retina abolished not only directional selectivity of ganglion cell responses but also an optokinetic eye reflex derived by stimulus movement. Starburst cells therefore serve as the key element that discriminates the direction of stimulus movement through integrative synaptic transmission and play a pivotal role in information processing that stabilizes image motion.

Activation of peroxisome proliferator-activated receptor gamma suppresses mast cell maturation involved in allergic diseases.

BACKGROUND: Mast cells play a central role in allergic and inflammatory diseases. Several reports indicated role of peroxisome proliferator-activated receptor gamma (PPARgamma) on mast cell function. However, there is no report about the role of PPARgamma on differentiation of mast cells from the progenitors. In this study, we investigated the role of PPARgamma in regulating bone marrow-derived mast cell maturation and the therapeutic implications for mast cell-related diseases such as atopic or contact dermatitis. METHODS: We used in vitro cell culture system for mast cell differentiation from bone marrow-progenitors using specific ligands and lentiviral-mediated short hairpin RNA of PPARgamma, and in vivo murine dermatitis models. RESULTS: Activation of PPARgamma inhibited the maturation of bone marrow progenitors into connective tissue-type mast cells (CTMCs) through up-regulation of GATA-4 and GATA-6 resulting in a decrease in expression of histidine decarboxylase and mast cell histamine content. In comparison, the differentiation of bone marrow progenitors into CTMCs was significantly accelerated by the knockdown of PPARgamma expression by lentiviral-mediated short hairpin RNA. Peroxisome proliferator-activated receptor gamma ligand administration to mice inhibited the maturation of mast cells resulting in attenuation of atopic and contact dermatitis via diminishment of the number of mature mast cells. CONCLUSION: Our results indicate that PPARgamma is one of master regulators on mast cell maturation and potentially useful for the therapy in various disorders involving mast cell activation.

Telomere maintenance in telomerase-deficient mouse embryonic stem cells: characterization of an amplified telomeric DNA.

Telomere dynamics, chromosomal instability, and cellular viability were studied in serial passages of mouse embryonic stem (ES) cells in which the telomerase RNA (mTER) gene was deleted. These cells lack detectable telomerase activity, and their growth rate was reduced after more than 300 divisions and almost zero after 450 cell divisions. After this growth crisis, survivor cells with a rapid growth rate did emerge. Such survivors were found to maintain functional telomeres in a telomerase-independent fashion. Although telomerase-independent telomere maintenance has been reported for some immortalized mammalian cells, its molecular mechanism has not been elucidated. Characterization of the telomeric structures in one of the survivor mTER(-/-) cell lines showed amplification of the same tandem arrays of telomeric and nontelomeric sequences at most of the chromosome ends. This evidence implicates cis/trans amplification as one mechanism for the telomerase-independent maintenance of telomeres in mammalian cells.

Expression and prognostic significance of EFNB2 and EphB4 genes in patients with oesophageal squamous cell carcinoma.

OBJECTIVE: Tyrosine kinases and its receptors play important roles in growth, migration, and invasion of malignant cells. Among those, there are only few reports examining the expression pattern of Eph/ephrin signalling system in oesophageal carcinoma. The prognostic importance of ephrin-B2 ligand (EFNB2) and its receptor EphB4, and its correlation with clinicopathologic characteristics are yet to be delineated in patients with oesophageal carcinoma. MATERIALS AND METHODS: EFNB2 gene and EphB4 receptor gene were examined of mRNA specimens in 61 patients with oesophageal squamous cell carcinoma using reverse-transcriptase polymerase chain reaction. EFNB2 protein was selectively examined using an immunohistochemical analysis. RESULTS: EFNB2 mRNA expression was detected in 38 (62.3%) and EphB4 expression was found in 44 (72.1%) out of 61 cancer tissues analysed. There was a statistically significant correlation between EFNB2 expression and number of lymph node metastasis (P<0.05), and a trend toward statistical significance for correlation between EFNB2 expression and American Joint Committee on Cancer Classification Stage (P<0.1), indicating that EFNB2 expression was up-regulated by advancement of the disease process. EFNB2 protein was strongly expressed in tumour with high mRNA EFNB2 expression and was weakly expressed in tumour with low mRNA expression in some representative tumours. The 5-year overall survival rate (23%) of patients with positive EFNB2 gene expression was significantly worse than 55% of negative expression (P<0.05). The results of multivariate analysis of prognosticators for survival showed that positive EFNB2 gene expression (P<0.01) and number of lymph node metastasis (P<0.05) were identified as significant factors indicative of a poorer survival. CONCLUSIONS: EFNB2 gene expression may be a biological marker and a useful prognostic indicator in patients with oesophageal squamous cell carcinoma.

Investigation of the influence of swallowing, coughing and vocalization on heart rate variability with respiratory-phase domain analysis.

OBJECTIVES: The objective of our study is to investigate extrinsic influences on heart rate variability using respiratory-phase domain analysis. Swallowing, coughing and vocalization (reading aloud and conversation) are adopted as extrinsic influences. METHODS: In this study, an instantaneous R-R interval (RRI) is sampled at each pi/10 rad of the respiratory phase and the data is divided into three subsets: a) respiration with event, b) one respiration after the event, and c) normal respiration. Then the mean waveforms of respiratory sinus arrhythmia (RSA) are calculated and compared. RESULTS AND CONCLUSIONS: It is found that swallowing induces tachycardia that recovers within one respiration. Coughing also induces tachycardia, but it does not recover within one respiration. Vocalization shortens the mean RRI, but the changing respiratory pattern due to vocalization has no statistically significant influence on the amplitude of RSA. Furthermore, it is found that the proposed method is effective for analyzing extrinsic influences on heart rate variability (HRV).