Unconventional miR-122 binding stabilizes the HCV genome by forming a trimolecular RNA structure.

MicroRNAs (miRNAs) typically downregulate protein expression from target mRNAs through limited base-pairing interactions between the 5' 'seed' region of the miRNA and the mRNA 3' untranslated region (3'UTR). In contrast to this established mode of action, the liver-specific human miR-122 binds at two sites within the hepatitis C viral (HCV) 5'UTR, leading to increased production of infectious virions. We show here that two copies of miR-122 interact with the HCV 5'UTR at partially overlapping positions near the 5' end of the viral transcript to form a stable ternary complex. Both miR-122 binding sites involve extensive base pairing outside of the seed sequence; yet, they have substantially different interaction affinities. Structural probing reveals changes in the architecture of the HCV 5'UTR that occur on interaction with miR-122. In contrast to previous reports, however, results using both the recombinant cytoplasmic exonuclease Xrn1 and liver cell extracts show that miR-122-mediated protection of the HCV RNA from degradation does not correlate with stimulation of viral propagation in vivo. Thus, the miR-122:HCV ternary complex likely functions at other steps critical to the viral life cycle

Anatomical Design and Production of a Novel 3-Dimensional Co-Culture System Replicating the Human Flexor Digitorum Profundus Enthesis

The enthesis, the specialized junction between tendon and bone, is a common site of injury. Although notoriously difficult to repair, advances in interfacial tissue engineering techniques are being developed for restorative function. Most notably are 3D in vitro co-culture models, built to recreate the complex heterogeneity of the native enthesis. While cell and matrix properties are often considered, there has been little attention given to native enthesis anatomical morphometrics and replicating these to enhance clinical relevance. This study focuses on the flexor digitorum profundus (FDP) tendon enthesis and, by combining anatomical morphometrics with computer-aided design, demonstrates the design and construction of an accurate and scalable model of the FDP enthesis. Bespoke 3D-printed mould inserts were fabricated based on the size, shape and insertion angle of the FDP enthesis. Then, silicone culture moulds were created, enabling the production of bespoke anatomical culture zones for an in vitro FDP enthesis model. The validity of the model has been confirmed using brushite cement scaffolds seeded with osteoblasts (bone) and fibrin hydrogel scaffolds seeded with fibroblasts (tendon) in individual studies with cells from either human or rat origin. This novel approach allows a bespoke anatomical design for enthesis repair and should be applied to future studies in this area.<br/

Two RNA-binding motifs in eIF3 direct HCV IRES-dependent translation.

The initiation of protein synthesis plays an essential regulatory role in human biology. At the center of the initiation pathway, the 13-subunit eukaryotic translation initiation factor 3 (eIF3) controls access of other initiation factors and mRNA to the ribosome by unknown mechanisms. Using electron microscopy (EM), bioinformatics and biochemical experiments, we identify two highly conserved RNA-binding motifs in eIF3 that direct translation initiation from the hepatitis C virus internal ribosome entry site (HCV IRES) RNA. Mutations in the RNA-binding motif of subunit eIF3a weaken eIF3 binding to the HCV IRES and the 40S ribosomal subunit, thereby suppressing eIF2-dependent recognition of the start codon. Mutations in the eIF3c RNA-binding motif also reduce 40S ribosomal subunit binding to eIF3, and inhibit eIF5B-dependent steps downstream of start codon recognition. These results provide the first connection between the structure of the central translation initiation factor eIF3 and recognition of the HCV genomic RNA start codon, molecular interactions that likely extend to the human transcriptome

Breaking up is hard to do : the economic impact of provisional funding contingent upon evidence development

Funding contingent upon evidence development (FED) has recently been the subject of some considerable debate in the literature but relatively little has been made of its economic impact. We argue that FED has the potential to shorten the lag between innovation and access but may also (i) crowd-out more valuable interventions in situations in which there is a fixed dedicated budget; or (ii) lead to a de facto increase in the funding threshold and increased expenditure growth in situations in which the programme budget is open-ended. Although FED would typically entail periodic review of provisional or interim listings, it may prove difficult to withdraw funding even at cost/QALY ratios well in excess of current listing thresholds. Further consideration of the design and implementation of FED processes is therefore required to ensure that its introduction yields net benefits over existing processes

Histomorphology of the subregions of the scapholunate ligament and its enthesis

Background  The scapholunate interosseous ligament (SLIL) has three subregions: dorsal, proximal, and volar. The SLIL enthesis has not previously been studied despite its important mechanical function in wrist joint biomechanics. Questions/Purposes  This study aims to compare the histomorphological differences between the SLIL subregions, including at their entheses. Three questions are explored: Do the gross dimensions differ between SLIL subregions? Does the enthesis qualitatively, and its calcified fibrocartilage (CF) quantitatively, differ between (a) SLIL subregions and (b) scaphoid and lunate attachments? Methods  Twelve fresh-frozen human cadaveric wrists were dissected and the gross dimensions of the SLIL subregions measured. Subregions were histologically processed for morphological and compositional analyses, including quantification of enthesis CF area. Results  The dorsal subregion was the thickest. The dorsal and volar subregions had fibrocartilaginous entheses, while the proximal subregion was attached to articular cartilage. The dorsal subregion had significantly more CF than the volar subregion. There was no significant difference in the enthesis CF between scaphoid and lunate attachments in the three subregions. Conclusions  There are significant morphological differences between the SLIL subregions. The dorsal subregion has the largest amount of CF, which is consistent with the greater biomechanical force subjected to this subregion. The similar histomorphology of the ligament at the scaphoid and lunate entheses suggests that similar biomechanical forces are applied to both attachments. Clinical Relevance  The histomorphological results confirm that the dorsal subregion is the strongest of the three subregions. The results from the entheseal region may have important implications in the study of graft incorporation during SLIL reconstruction

Design and Development of a Bioreactor System for Mechanical Stimulation of Musculoskeletal Tissue

We report on the development of a bioreactor system for mechanical stimulation of musculoskeletal tissues. The ultimate object is to improve the quality of medical treatment following injuries of the enthesis tissue. To this end, the tissue formation process through the effect of mechanical stimulation is investigated. A six-well system was designed, 3D printed and tested. An integrated actuator creates strain by applying a force. A contactless position sensor monitors the travels. An electronic circuit controls the bioreactor using a microcontroller. An IoT platform connects the microcontroller to a smartphone, enabling the user to alter variables, trigger actions and monitor the system. The system was stabilised by implementing two PID controllers and safety measures. The results show that the bioreactor design is suited to execute mechanical stimulation and to investigate the tissue formation and regeneration process. The bioreactor reported here can now be implemented in tissue engineering applications including tissue specimen.</p

Innominate Artery Pseudoaneurysm From a Salter-Harris Fracture of the Sternoclavicular Joint

Fractures and dislocations of the sternoclavicular joint (SCJ) are uncommon, accounting for \u3c5% of all shoulder girdle injuries. They are relatively more common in the pediatric population than in the adult population and can often present concurrently as a posteriorly displaced medial clavicular dislocation with a fracture through the unfused physis. It is especially important to recognize this injury, because its management and potential sequelae are very different from those for fractures of the clavicle shaft. This type of injury frequently requires closed or open operative management because fracture-dislocation of the SCJ can be associated with potentially serious complications such as pneumothorax, brachial plexus injury, vagus nerve injury, tracheal injury, and vascular compromise. Few case reports describe fracture-dislocation of the SCJ resulting in vascular injuries. We describe the case of a 17-year-old boy who sustained a blunt hockey injury resulting in a right physeal fracture-dislocation of the SCJ causing an innominate artery pseudoaneurysm. This was treated with excision of the pseudoaneurysm, bovine pericardial patch angioplasty repair of the innominate artery, and open reduction and internal fixation of the medial clavicular physeal fracture

Lignin biosynthesis perturbations affect secondary cell wall composition and saccharification yield in Arabidopsis thaliana.

BACKGROUND: Second-generation biofuels are generally produced from the polysaccharides in the lignocellulosic plant biomass, mainly cellulose. However, because cellulose is embedded in a matrix of other polysaccharides and lignin, its hydrolysis into the fermentable glucose is hampered. The senesced inflorescence stems of a set of 20 Arabidopsis thaliana mutants in 10 different genes of the lignin biosynthetic pathway were analyzed for cell wall composition and saccharification yield. Saccharification models were built to elucidate which cell wall parameters played a role in cell wall recalcitrance. RESULTS: Although lignin is a key polymer providing the strength necessary for the plant's ability to grow upward, a reduction in lignin content down to 64% of the wild-type level in Arabidopsis was tolerated without any obvious growth penalty. In contrast to common perception, we found that a reduction in lignin was not compensated for by an increase in cellulose, but rather by an increase in matrix polysaccharides. In most lignin mutants, the saccharification yield was improved by up to 88% cellulose conversion for the cinnamoyl-coenzyme A reductase1 mutants under pretreatment conditions, whereas the wild-type cellulose conversion only reached 18%. The saccharification models and Pearson correlation matrix revealed that the lignin content was the main factor determining the saccharification yield. However, also lignin composition, matrix polysaccharide content and composition, and, especially, the xylose, galactose, and arabinose contents influenced the saccharification yield. Strikingly, cellulose content did not significantly affect saccharification yield. CONCLUSIONS: Although the lignin content had the main effect on saccharification, also other cell wall factors could be engineered to potentially increase the cell wall processability, such as the galactose content. Our results contribute to a better understanding of the effect of lignin perturbations on plant cell wall composition and its influence on saccharification yield, and provide new potential targets for genetic improvement.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Characterising the nicotine metabolite ratio and its association with treatment choice: A cross sectional analysis of Stop Smoking Services in England

Pharmacotherapy provision based on Nicotine Metabolite Ratio (NMR) status (slow/normal metabolism) may improve smoking cessation rates. However, it is unclear whether NMR status is consistent across patient characteristics and current treatment choice. Data come from 1,826 participants attending Stop Smoking Services (SSS) across England in 2012/13. Sociodemographic, mental/physical health, smoking and treatment characteristics (nicotine replacement therapy vs. other pharmacotherapy; group vs. one-to-one behavioural support) were assessed. Salivary nicotine metabolites were measured and NMR (3-hydroxycotinine/cotinine) computed, characterising smokers as slow (NMR < 0.31) or normal (NMR ≥ 0.31) metabolisers. Normal metabolisers were older than slow metabolisers (Odds Ratio (OR) = 1.49, 95% Confidence Interval (CI) = 1.32-1.69) but no other characteristics were associated with NMR status. Overall, predictors accounted for only 7.3% of NMR variance. In adjusted analysis, pharmacotherapy type was not associated with NMR status, but normal metabolisers were less likely to use group support (OR = 0.67, 95% CI = 0.51-0.89). NMR status does not vary substantially across sociodemographic characteristics. Given its impact on pharmacotherapy efficacy, the lack of an association with pharmacotherapy choice suggests there is scope to use NMR status to optimise the selection and efficacy of smoking cessation pharmacotherapy. The unexpected association of NMR status with behavioural support should be explored further