TXNIP in liver sinusoidal endothelial cells ameliorates alcohol-associated liver disease via nitric oxide production.

Dysregulation of liver sinusoidal endothelial cell (LSEC) differentiation and function has been reported in alcohol-associated liver disease (ALD). Impaired nitric oxide (NO) production stimulates LSEC capillarization and dysfunction; however, the mechanism underlying NO production remains unclear. Here, we investigated the role of thioredoxin-interacting protein (TXNIP), an important regulator of redox homeostasis, in endothelial cell NO production and its subsequent effects on ALD progression. We found that hepatic TXNIP expression was upregulated in patients with ALD and in ethanol diet-fed mice with high expression in LSECs. Endothelial cell-specific Txnip deficiency (TxnipΔEC) in mice exacerbated alcohol-induced liver injury, inflammation, fibrosis, and hepatocellular carcinoma development. Deletion of Txnip in LSECs led to sinusoidal capillarization, downregulation of NO production, and increased release of proinflammatory cytokines and adhesion molecules, whereas TXNIP overexpression had the opposite effects. Mechanistically, TXNIP interacted with transforming growth factor ß-activated kinase 1 (TAK1) and subsequently suppressed the TAK1 pathway. Inhibition of TAK1 activation restored NO production and decreased the levels of proinflammatory cytokines, thereby, blocking liver injury and inflammation in TxnipΔEC mice. Our findings indicate that upregulated TXNIP expression in LSECs serves a protective role in ameliorating ALD. Enhancing TXNIP expression could, therefore, be a potential therapeutic approach for ALD.

Selective Inhibition of ADAR1 Using 8-Azanebularine-Modified RNA Duplexes.

Adenosine deaminases acting on RNA (ADARs) are RNA editing enzymes that catalyze the hydrolytic deamination of adenosine (A) to inosine (I) in dsRNA. In humans, two catalytically active ADARs, ADAR1 and ADAR2, perform this A-to-I editing event. The growing field of nucleotide base editing has highlighted ADARs as promising therapeutic agents while multiple studies have also identified ADAR1's role in cancer progression. However, the potential for site-directed RNA editing as well as the rational design of inhibitors is being hindered by the lack of detailed molecular understanding of RNA recognition by ADAR1. Here, we designed short RNA duplexes containing the nucleoside analog, 8-azanebularine (8-azaN), to gain insight into molecular recognition by the human ADAR1 catalytic domain. From gel shift and in vitro deamination experiments, we validate ADAR1 catalytic domain's duplex secondary structure requirement and present a minimum duplex length for binding (14 bp, with 5 bp 5' and 8 bp 3' to editing site). These findings concur with predicted RNA-binding contacts from a previous structural model of the ADAR1 catalytic domain. Finally, we establish that neither 8-azaN as a free nucleoside nor a ssRNA bearing 8-azaN inhibits ADAR1 and demonstrate that the 8-azaN-modified RNA duplexes selectively inhibit ADAR1 and not the closely related ADAR2 enzyme.

Patient discomfort levels during instrumentation procedure using nickel-titanium files with different kinetic movements.

This study evaluated the perceived vibration, noise and discomfort levels associated with two nickel-titanium file systems with different kinetics; reciprocating motion (REC) using WaveOne Gold and continuous rotation motion (CON) using ProTaper NEXT. Forty roots with two canals from maxillary premolar and molar of 40 patients were included. Root canals were instrumented using each system for each canal. Patients were surveyed about the vibration, noise and discomfort experienced using visual analogue scale, and their preference. The responses were statistically analysed using Wilcoxon Signed-Rank test, Mann-Whitney U test and Spearman's rank correlation test at the 95% of significance level. The vibration, noise and discomfort experienced were significantly greater in REC than CON (P < 0.05). In REC, male subjects reported significantly higher vibration than female (P < 0.05). Majority respondents (72.5%) preferred the CON method. The perceived vibration, noise and discomfort were less apparent from the CON than the REC.

Scanning transmission X-ray microscopy study of subcellular granules in human platelets at the carbon K- and calcium L2,3-edges.

Platelets and their subcellular components (e.g., dense granules) are essential components in hemostasis. Understanding their chemical heterogeneities at the sub-micrometer scale, particularly their activation during hemostasis and production of platelet-derived extracellular vesicles, may provide important insights into their mechanisms; however, this has rarely been investigated, mainly owing to the lack of appropriate chemical characterization tools at nanometer scale. Here, the use of scanning transmission X-ray microscopy (STXM) combined with X-ray absorption near edge structure (XANES) to characterize human platelets and their subcellular components at the carbon K-edge and calcium L2,3-edge, is reported. STXM images can identify not only the spatial distribution of subcellular components in human platelets, such as dense granules (DGs) with sizes of ~200 nm, but also their granule-to-granule chemical heterogeneities on the sub-micrometer scale, based on their XANES spectra. The calcium distribution map as well as the principal component analysis of the STXM image stacks clearly identified the numbers and locations of the calcium-rich DGs within human platelets. Deconvolution of the carbon K-edge XANES spectra, extracted from various locations in the platelets, showed that amide carbonyl and carboxylic acid functional groups were mainly found in the cytoplasm, while ketone-phenol-nitrile-imine, aliphatic, and carbonate functional groups were dominant in the platelet DGs. These observations suggest that platelet DGs are most likely composed of calcium polyphosphate associated with adenosine triphosphate (ATP) and adenosine diphosphate (ADP), with significant granule-to-granule variations in their compositions, while the cytoplasm regions of platelets contain significant amounts of proteins.

High-throughput mutagenesis reveals unique structural features of human ADAR1.

Adenosine Deaminases that act on RNA (ADARs) are enzymes that catalyze adenosine to inosine conversion in dsRNA, a common form of RNA editing. Mutations in the human ADAR1 gene are known to cause disease and recent studies have identified ADAR1 as a potential therapeutic target for a subset of cancers. However, efforts to define the mechanistic effects for disease associated ADAR1 mutations and the rational design of ADAR1 inhibitors are limited by a lack of structural information. Here, we describe the combination of high throughput mutagenesis screening studies, biochemical characterization and Rosetta-based structure modeling to identify unique features of ADAR1. Importantly, these studies reveal a previously unknown zinc-binding site on the surface of the ADAR1 deaminase domain which is important for ADAR1 editing activity. Furthermore, we present structural models that explain known properties of this enzyme and make predictions about the role of specific residues in a surface loop unique to ADAR1.

Asymmetric dimerization of adenosine deaminase acting on RNA facilitates substrate recognition.

Adenosine deaminases acting on RNA (ADARs) are enzymes that convert adenosine to inosine in duplex RNA, a modification that exhibits a multitude of effects on RNA structure and function. Recent studies have identified ADAR1 as a potential cancer therapeutic target. ADARs are also important in the development of directed RNA editing therapeutics. A comprehensive understanding of the molecular mechanism of the ADAR reaction will advance efforts to develop ADAR inhibitors and new tools for directed RNA editing. Here we report the X-ray crystal structure of a fragment of human ADAR2 comprising its deaminase domain and double stranded RNA binding domain 2 (dsRBD2) bound to an RNA duplex as an asymmetric homodimer. We identified a highly conserved ADAR dimerization interface and validated the importance of these sequence elements on dimer formation via gel mobility shift assays and size exclusion chromatography. We also show that mutation in the dimerization interface inhibits editing in an RNA substrate-dependent manner for both ADAR1 and ADAR2.

Off-Target Editing by CRISPR-Guided DNA Base Editors.

Base editing is a genome editing strategy that induces specific single-nucleotide changes within genomic DNA. Two major DNA base editors, cytosine base editors and adenine base editors, that consist of a Cas9 protein linked to a deaminase enzyme that catalyzes targeted base conversion directed by a single-guide RNA have been developed. This strategy has been used widely for precise genome editing because, unlike CRISPR-Cas nuclease-based genome editing systems, this strategy does not create double-strand DNA breaks that often result in high levels of undesirable indels. However, recent papers have reported that DNA base editors can cause substantial off-target editing in both genomic DNA and RNA. The off-target editing described in these studies is primarily independent of guide RNA and arises from the promiscuous reactivity of the deaminase enzymes used in DNA base editors. In this Perspective, we discuss the development of DNA base editors, the guide RNA-independent off-target activity reported in recent studies, and strategies that improve the selectivity of DNA base editors.

Salinomycin Inhibits Influenza Virus Infection by Disrupting Endosomal Acidification and Viral Matrix Protein 2 Function.

Screening of chemical libraries with 2,000 synthetic compounds identified salinomycin as a hit against influenza A and B viruses, with 50% effective concentrations ranging from 0.4 to 4.3 µM in cells. This compound is a carboxylic polyether ionophore that exchanges monovalent ions for protons across lipid bilayer membranes. Monitoring the time course of viral infection showed that salinomycin blocked nuclear migration of viral nuclear protein (NP), the most abundant component of the viral ribonucleoprotein (vRNP) complex. It caused cytoplasmic accumulation of NP, particularly within perinuclear endosomes, during virus entry. This was primarily associated with failure to acidify the endosomal-lysosomal compartments. Similar to the case with amantadine (AMT), proton channel activity of viral matrix protein 2 (M2) was blocked by salinomycin. Using purified retroviral Gag-based virus-like particles (VLPs) with M2, it was proved that salinomycin directly affects the kinetics of a proton influx into the particles but in a manner different from that of AMT. Notably, oral administration of salinomycin together with the neuraminidase inhibitor oseltamivir phosphate (OSV-P) led to enhanced antiviral effect over that with either compound used alone in influenza A virus-infected mouse models. These results provide a new paradigm for developing antivirals and their combination therapy that control both host and viral factors.IMPORTANCE Influenza virus is a main cause of viral respiratory infection in humans as well as animals, occasionally with high mortality. Circulation of influenza viruses resistant to the matrix protein 2 (M2) inhibitor, amantadine, is highly prevalent. Moreover, the frequency of detection of viruses resistant to the neuraminidase inhibitors, including oseltamivir phosphate (OSV-P) or zanamivir, is also increasing. These issues highlight the need for discovery of new antiviral agents with different mechanisms. Salinomycin as the monovalent cation-proton antiporter exhibited consistent inhibitory effects against influenza A and B viruses. It plays multifunctional roles by blocking endosomal acidification and by inactivating the proton transport function of M2, the key steps for influenza virus uncoating. Notably, salinomycin resulted in marked therapeutic effects in influenza virus-infected mice when combined with OSV-P, suggesting that its chemical derivatives could be developed as an adjuvant antiviral therapy to treat influenza infections resistant or less sensitive to existing drugs.

Selective Recognition of RNA Substrates by ADAR Deaminase Domains.

Adenosine deamination is one of the most prevalent post-transcriptional modifications in mRNA and is catalyzed by ADAR1 and ADAR2 in humans. ADAR1 and ADAR2 have different substrate selectivity, which is believed to mainly originate from the proteins' deaminase domains (hADAR1d and hADAR2d, respectively). RNA-seq of the Saccharomyces cerevisiae transcriptome subjected to ADAR-catalyzed RNA editing identified substrates with common secondary structure features preferentially edited by hADAR1d over hADAR2d. The relatively small size and efficient reaction of one of these substrates suggested it could be useful for further study of the hADAR1d reaction. Indeed, a short hairpin stem from the S. cerevisiae HER1 mRNA was efficiently deaminated by hADAR1d and used to generate an hADAR1d-specific fluorescent reporter of editing activity. Using substrates preferred by either hADAR1d or hADAR2d in vitro, we found that a chimeric protein bearing an RNA-binding loop from hADAR2d grafted onto hADAR1d showed ADAR2-like selectivity. Finally, a high-throughput mutagenesis analysis (Sat-FACS-Seq) of conserved residues in an RNA-binding loop of hADAR1d revealed essential amino acids for function, advancing our understanding of RNA recognition by this domain.

Evolutionary relationships of the hexon and penton base genes of novel squirrel adenovirus.

Squirrel adenovirus (SqAdV) was reported previously. However, only partial sequences of its hexon and polymerase genes have been revealed. For the first time, we report the full-length genome of SqAdV including the complete hexon and penton base genes. From internal body organs of 59 red squirrels archived in Korea Bank for Pathogenic Viruses, the hexon, penton base, and full-length genome of SqAdV were determined by a PCR method. Of the internal body organs examined, the spleen showed the highest detection rate (25.42%) for SqAdV whereas the kidney and lung exhibited 18.64% and 3.39% rates, respectively. Based on the phylogenetic relationships of the hexon and penton base genes, SqAdV appears to belong to the genus Mastadenovirus, and, at least in our study, the hexon of SqAdV exhibits the closest relationship to that of an alpaca AdV. Compared with the hexon, the penton base of SqAdV appears to be genetically more divergent from that of other mastadenoviruses. It was also revealed that the full-length SqAdV genome retained AT nucleotide content similar level to AT-rich atadenoviruses, which is unusual for mastadenoviruses. Our results emphasize that SqAdV is classified into the genus Mastadenovirus and demonstrate the AT-biased nucleotide constitution of SqAdV.

Differential diagnosis of periapical cyst using collagen birefringence pattern of the cyst wall.

OBJECTIVES: Periapical lesions, including periapical cyst (PC), periapical granuloma (PG), and periapical abscess (PA), are frequently affected by chemical/physical damage during root canal treatment or severe bacterial infection, and thus, the differential diagnosis of periapical lesions may be difficult due to the presence of severe inflammatory reaction. The aim of this study was to make differential diagnosis among PC, PG, and PA under polarizing microscope. MATERIALS AND METHODS: The collagen birefringence patterns of 319 cases of PC (n = 122), PG (n = 158), and PA (n = 39) obtained using a polarizing microscope were compared. In addition, 6 cases of periodontal fibroma (PF) were used as positive controls. RESULTS: Collagen birefringence was condensed with a thick, linear band-like pattern in PC, but was short and irregularly scattered in PG, and scarce or absent in PA. PF showed intense collagen birefringence with a short, palisading pattern but no continuous band-like pattern. The linear band-like birefringence in PC was ascribed to pre-existing expansile tensile stress of the cyst wall. CONCLUSIONS: In this study all PCs (n = 122) were distinguishable from PGs and PAs by their characteristic birefringence, despite the absence of lining epithelium (n = 20). Therefore, the authors suggest that the presence of linear band-like collagen birefringence of the cyst wall aids the diagnostic differentiation of PC from PG and PA.

Vibrations Generated by Several Nickel-titanium Endodontic File Systems during Canal Shaping in an Ex Vivo Model.

INTRODUCTION: This study aimed to compare the vibration generated by several nickel-titanium (NiTi) file systems and transmitted to teeth under 2 different motions (continuous rotation motion and reciprocating motion). METHODS: Sixty J-shaped resin blocks (Endo Training Bloc-J; Dentsply Maillefer, Ballaigues, Switzerland) were trimmed to a root-shaped form and divided into 2 groups according to the types of electric motors: WaveOne motor (WOM, Dentsply Maillefer) and X-Smart Plus motor (XSM, Dentsply Maillefer). Each group was further subdivided into 3 subgroups (n = 10 each) according to the designated file systems: ProTaper Next (PTN, Dentsply Maillefer), ProTaper Universal (PTU, Dentsply Maillefer), and WaveOne (WOP, Dentsply Maillefer) systems. Vibration was measured during the pecking motion using an accelerometer attached to a predetermined consistent position. The average vibration values were subjected to 2-way analysis of variance as well as the t test and Duncan test for post hoc comparison at the 95% confidence interval. RESULTS: Both motor types and instrument types produced significantly different ranges of average vibrations. Regardless of the instrument types, the WOM group generated greater vibration than the XSM group (P < .05). Although PTN and PTU did not show significant differences, the WOP group showed significantly greater vibration than the other groups regardless of motor types (P < .05). CONCLUSIONS: Under the limitations of this study design, the reciprocating NiTi file system may generate greater vibration than the continuous rotation NiTi file systems. The motor type also has a significant effect to amplify the vibrations.

The geometric effect of an off-centered cross-section on nickel-titanium rotary instruments: A finite element analysis study.

BACKGROUND/PURPOSE: Geometric design dictates the mechanical performance of nickel-titanium rotary instruments. Using finite element (FE) analysis, this study evaluated the effects of an off-centered cross-sectional design on the stiffness and stress distribution of nickel-titanium rotary instruments. MATERIALS AND METHODS: We constructed three-dimensional FE models, using ProTaper-NEXT type design (PTN) as well as three other virtual instruments with varied cross-sectional aspect ratios but all with the same cross-sectional area. The cross-sectional aspect ratio of the PTN was 0.75, while others were assigned to have ratios of 1.0 (square), 1.5 (rectangle), and 2.215 (centered-rectangle). The PTN center of the cross-section was 'k', while others were designed to have 0.9992k, 0.7k, and 0 for the square, rectangle, and centered-rectangle models, respectively. To compare the stiffness of the four FE models, we numerically analyzed their mechanical response under bending and torque. RESULTS: Under the bending condition, the square model was found to be the stiffest, followed by the PTN, rectangle, and then the centered-rectangle model. Under the torsion, the square model had the smallest distortion angle, while the rectangular model had the highest distortion angle. CONCLUSION: Under the limitation of this study, the PTN type off-centered cross-sectional design appeared the most optimal configuration among the tested designs for high bending stiffness with cutting efficiency while rotational stiffness remained similar with the other designs.

Increased sclerostin levels after further ablation of remnant estrogen by aromatase inhibitors.

BACKGROUND: Sclerostin is a secreted Wnt inhibitor produced almost exclusively by osteocytes, which inhibits bone formation. Aromatase inhibitors (AIs), which reduce the conversion of steroids to estrogen, are used to treat endocrine-responsive breast cancer. As AIs lower estrogen levels, they increase bone turnover and lower bone mass. We analyzed changes in serum sclerostin levels in Korean women with breast cancer who were treated with an AI. METHODS: We included postmenopausal women with endocrine-responsive breast cancer (n=90; mean age, 57.7 years) treated with an AI, and compared them to healthy premenopausal women (n=36; mean age, 28.0 years). The subjects were randomly assigned to take either 5 mg alendronate with 0.5 µg calcitriol (n=46), or placebo (n=44) for 6 months. RESULTS: Postmenopausal women with breast cancer had significantly higher sclerostin levels compared to those in premenopausal women (27.8±13.6 pmol/L vs. 23.1±4.8 pmol/L, P<0.05). Baseline sclerostin levels positively correlated with either lumbar spine or total hip bone mineral density only in postmenopausal women (r=0.218 and r=0.233; P<0.05, respectively). Serum sclerostin levels increased by 39.9%±10.2% 6 months after AI use in postmenopausal women; however, no difference was observed between the alendronate and placebo groups (39.9%±10.2% vs. 55.9%±9.13%, P>0.05). CONCLUSION: Serum sclerostin levels increased with absolute deficiency of residual estrogens in postmenopausal women with endocrine-responsive breast cancer who underwent AI therapy with concurrent bone loss.

Combination effects of peramivir and favipiravir against oseltamivir-resistant 2009 pandemic influenza A(H1N1) infection in mice.

Antiviral drugs are being used for therapeutic purposes against influenza illness in humans. However, antiviral-resistant variants often nullify the effectiveness of antivirals. Combined medications, as seen in the treatment of cancers and other infectious diseases, have been suggested as an option for the control of antiviral-resistant influenza viruses. Here, we evaluated the therapeutic value of combination therapy against oseltamivir-resistant 2009 pandemic influenza H1N1 virus infection in DBA/2 mice. Mice were treated for five days with favipiravir and peramivir starting 4 hours after lethal challenge. Compared with either monotherapy, combination therapy saved more mice from viral lethality and resulted in increased antiviral efficacy in the lungs of infected mice. Furthermore, the synergism between the two antivirals, which was consistent with the survival outcomes of combination therapy, indicated that favipiravir could serve as a critical agent of combination therapy for the control of oseltamivir-resistant strains. Our results provide new insight into the feasibility of favipiravir in combination therapy against oseltamivir-resistant influenza virus infection.

Spirosoma radiotolerans sp. nov., a gamma-radiation-resistant bacterium isolated from gamma ray-irradiated soil.

A Gram-negative, short-rod-shaped bacterial strain with gliding motility, designated as DG5A(T), was isolated from a rice field soil in South Korea. Phylogenic analysis using 16S rRNA gene sequence of the new isolate showed that strain DG5A(T) belong to the genus Spirosoma in the family Spirosomaceae, and the highest sequence similarities were 95.5 % with Spirosoma linguale DSM 74(T), 93.4 % with Spirosoma rigui WPCB118(T), 92.8 % with Spirosoma luteum SPM-10(T), 92.7 % with Spirosoma spitsbergense SPM-9(T), and 91.9 % with Spirosoma panaciterrae Gsoil 1519(T). Strain DG5A(T) revealed resistance to gamma and UV radiation. Chemotaxonomic data showed that the most abundant fatty acids were summed feature C(16:1) ω7c/C(16:1) ω6c (36.90 %), C(16:1) ω5c (29.55 %), and iso-C(15:0) (14.78 %), and the major polar lipid was phosphatidylethanolamine (PE). The DNA G+C content of strain DG5A(T) was 49.1 mol%. Together, the phenotypic, phylogenetic, and chemotaxonomic data supported that strain DG5A(T) presents a novel species of the genus Spirosoma, for which the name Spirosoma radiotolerans sp. nov., is proposed. The type strain is DG5A(T) (=KCTC 32455(T) = JCM19447(T)).

A comparison of dimensional standard of several nickel-titanium rotary files.

OBJECTIVES: The aim of this study was to compare the dimensional standard of several nickel-titanium (Ni-Ti) rotary files and verify the size conformity. MATERIALS AND METHODS: ProFile (Dentsply Maillefer), RaCe (FKG Dentaire), and TF file (SybronEndo) #25 with a 0.04 and 0.06 taper were investigated, with 10 in each group for a total of 60 files. Digital images of Ni-Ti files were captured under light microscope (SZX16, Olympus) at 32×. Taper and diameter at D1 to D16 of each files were calculated digitally with AnalySIS TS Materials (OLYMPUS Soft Imaging Solutions). Differences in taper, the diameter of each level (D1 to D16) at 1 mm interval from (ANSI/ADA) specification No. 101 were statistically analyzed using one-way ANOVA and Scheffe's post-hoc test at 95% confidence level. RESULTS: TF was the only group not conform to the nominal taper in both tapers (p < 0.05). All groups except 0.06 taper ProFile showed significant difference from the nominal diameter (p < 0.05). CONCLUSIONS: Actual size of Ni-Ti file, especially TF, was different from the manufacturer's statements.

Chronic maxillary sinusitis caused by root canal overfilling of Calcipex II.

This is a case report of chronic maxillary sinusitis caused by root canal overfilling of Calcipex II (Techno-Dent). A 60 year-old male complained of dull pain in the right maxillary molar area after complicated endodontic treatment using Calcipex II paste and was finally diagnosed with a chronic maxillary sinusitis through a clinical and radiological observation. In the biopsy examination, the periapical granuloma contained a lot of dark and translucent Calcipex II granules which were not stained with hematoxylin and eosin. They were usually engulfed by macrophages but rarely resorbed, resulting in scattering and migrating into antral mucosa. Most of the Calcipex II granules were also accumulated in the cytoplasms of secretory columnar epithelial cells, and small amount of Calcipex II granules were gradually secreted into sinus lumen by exocytosis. However, chronic granulomatous inflammation occurred without the additional recruitment of polymorphonuclear leukocytes (PMNs) and lymphocytes, and many macrophages which engulfed the Calcipex II granules were finally destroyed in the processes of cellular apoptosis. It is presumed that Calcipex II granules are likely to have a causative role to induce the granulomatous foreign body inflammation in the periapical region, and subsequently to exacerbate the chronic maxillary sinusitis in this study.

Effect of torsional loading of nickel-titanium instruments on cyclic fatigue resistance.

INTRODUCTION: The aim of this study was to evaluate the effect of torsional preload on the cyclic fatigue life of nickel-titanium rotary instruments. METHODS: ProFile (#25/0.06) (Dentsply Maillefer, Ballaigues, Switzerland) and ProTaper (F1; Dentsply Maillefer) were used. Each file was preloaded at 4 conditions (ie, no preloading and 25%, 50%, and 75% of mean ultimate torsional strength) of torsional prestress before the fatigue test. The torsional preloads were applied by securing 5 mm of the file tip while keeping the file straight, rotating it clockwise until the preset torque, and then returning to the original position. This motion was repeated until a preset number (10, 30, or 50) of repetitions were reached (n = 12). After torsional preloading, the number of cycles to failure was evaluated in a simulated canal. Data were analyzed using 2-way analysis of variance and the Duncan post hoc comparison. The fractured fragment surfaces were examined under a scanning electron microscope for the topographic features of fractured instruments. RESULTS: For both instruments, there was a significant effect because of the extent of torsional preloads. The 50% and 75% torsionally preloaded ProFile and all ProTaper preloading groups had a higher number of cycles to failure than the other group(s). There was little difference in the lateral view appearance between new and torsionally preloaded files. After cyclic fatigue testing, all preloaded instruments showed numerous microcracks adjacent to the fracture site on lateral view examination. The microcracks did not seem to follow the machining grooves on the instrument surface but rather ran irregularly. CONCLUSIONS: The torsional preloads within the superelastic limit of the material may improve the cyclic fatigue resistance of nickel-titanium rotary instruments.

Aronia melanocarpa and its components demonstrate antiviral activity against influenza viruses.

The influenza virus is highly contagious in human populations around the world and results in approximately 250,000-500,000 deaths annually. Vaccines and antiviral drugs are commonly used to protect susceptible individuals. However, the antigenic mismatch of vaccines and the emergence of resistant strains against the currently available antiviral drugs have generated an urgent necessity to develop a novel broad-spectrum anti-influenza agent. Here we report that Aronia melanocarpa (black chokeberry, Aronia), the fruit of a perennial shrub species that contains several polyphenolic constituents, possesses in vitro and in vivo efficacy against different subtypes of influenza viruses including an oseltamivir-resistant strain. These anti-influenza properties of Aronia were attributed to two constituents, ellagic acid and myricetin. In an in vivo therapeutic mouse model, Aronia, ellagic acid, and myricetin protected mice against lethal challenge. Based on these results, we suggest that Aronia is a valuable source for antiviral agents and that ellagic acid and myricetin have potential as influenza therapeutics.