Molecular rulers for measuring RNA structure: sites of crosslinking in chlorambucilyl-phenylalanyl-tRNA^(Phe) (yeast) and chlorambucilyl-pentadecaprolyl-phenylalanyl-tRNA^(Phe) (yeast) intramolecularly crosslinked in aqueous solution

Intramolecular crosslinking of yeast phenylalanine tRNA in aqueous solution with rigid, variable-length crosslinking reagents, which we call "molecular rulers," has given results in reasonable agreement with the crystal structure. Chlorambucilyl-[^(3)H]phenylalanyl-tRNA^(Phe) crosslinked intramolecularly at G-71 and A-73, whereas chlorambucilyl-pentadecaprolyl-[^(3)H]phenylalanyl-tRNA^(Phe) crosslinked at G-20 and Y-37. The pentadecaprolyl reagent was predicted to be 62 Å long, including chlorambucil and phenylalanine; the sites that it reached are 60 Å distant from the 3' OH (in the case of G-20) or 80 Å distant (in the case of Y-37) in the crystal structure of tRNA^(Phe). The close agreement between the length of the reagent and the distance of G-20 from the 3' OH in the crystal structure illustrates the rigidity of the tRNA^(Phe) molecule in the dihydrouridine loop region at the corner of the molecule. The apparent ability of the 62-Å-long reagent to crosslink to a site, Y-37, that is 80 Å distant from the 3' OH in the crystal structure appears to illustrate the flexibility of both the 3' A-C-C-A terminus and the anticodon stem and loop, with respect to the tRNA molecule. These observations demonstrate the utility of oligoproline-based crosslinking reagents as rigid, variable-length molecular rulers for biological macromolecules in solution

Atypical periosteal osteoid osteoma: a case report

Osteoid osteoma is a benign osteoblastic tumor usually seen in adolescent and young males. In the paediatric age group, since the history may be difficult to elicit, there are often problems in early diagnosis. The author reports an unusual presentation of osteoid osteoma in a ten-year-old girl, which could not be diagnosed by conventional X-rays and CT scan

Coordinate control of cell cycle regulatory genes in zebrafish development tested by cyclin D1 knockdown with morpholino phosphorodiamidates and hydroxyprolyl-phosphono peptide nucleic acids

During early zebrafish (Danio rerio) development zygotic transcription does not begin until the mid-blastula transition (MBT) ∼3 h after fertilization. MBT demarcates transition from synchronous short cell cycles of S and M phases exclusively to full cycles encompassing G(1) and G(2) phases. Transcriptional profiling and RT–PCR analyses during these phases enabled us to determine that this shift corresponds to decreased transcript levels of S/M phase cell cycle control genes (e.g. ccna2, ccnb1, ccnb2 and ccne) and increased transcript levels of ccnd1, encoding cyclin D1, and orthologs of p21 (p21-like) and retinoblastoma (Rb-like 1). To investigate the regulation of this process further, the translation of ccnd1 mRNA, a G(1)/S checkpoint control element, was impaired by microinjection of ccnd1-specific morpholino phosphorodiamidate (MO) 20mer or hydroxyprolyl-phosphono peptide nucleic acid (HypNA-pPNA) 16mer antisense oligonucleotides. The resulting downregulation of cyclin D1 protein resulted in microophthalmia and microcephaly, but not lethality. The phenotypes were not seen with 3-mismatch MO 20mers or 1-mismatch HypNA-pPNA 16mers, and were rescued by an exogenous ccnd1 mRNA construct with five mismatches. Collectively, these results indicate that transcription of key molecular determinants of asynchronous cell cycle control in zebrafish embryos commences at MBT and that the reduction of cyclin D1 expression compromises zebrafish eye and head development

The inhibition of \u3cem\u3eStaphylococcus epidermidis\u3c/em\u3e biofilm formation by vancomycinmodified titanium alloy and implications for the treatment of periprosthetic infection

Peri-prosthetic infections are notoriously difficult to treat as the biomaterial implant is ideal for bacterial adhesion and biofilm formation, resulting in decreased antibiotic sensitivity. Previously, we reported that vancomycin covalently attached to a Ti alloy surface (Vanc-Ti) could prevent bacterial colonization. Herein we examine the effect of this Vanc-Ti surface on Staphylococci epidermidis, a Gram-positive organism prevalent in orthopaedic infections. By direct colony counting and fluorescent visualization of live bacteria, S. epidermidis colonization was significantly inhibited on Vanc-Ti implants. In contrast, the gram-negative organism Escherichia coli readily colonized the Vanc-Ti rod, suggesting retention of antibiotic specificity. By histochemical and SEM analysis, Vanc-Ti prevented S. epidermidis biofilm formation, even in the presence of serum. Furthermore, when challenged multiple times with S. epidermidis, Vanc-Ti rods resisted bacterial colonization. Finally, when S. epidermidis was continuously cultured in the presence of Vanc-Ti, the bacteria maintained a Vanc sensitivity equivalent to the parent strain. These findings indicate that antibiotic derivatization of implants can result in a surface that can resist bacterial colonization. This technology holds great promise for the prevention and treatment of periprosthetic infections

General practitioners apply the usual care for shoulder complaints better than expected – analysis of videotaped consultations

BACKGROUND: The education and activation program (EAP) is a newly developed intervention to prevent the development of chronic shoulder complaints (SCs). Trained general practitioners (GPs) administer the EAP. The EAP addresses inadequate cognitions and maladaptive behavior related to the SCs. The effect of the EAP is evaluated in a randomized clinical trial. The aim of the present study is to use videotaped consultations to study (1) the performance of trained GPs administering the EAP and (2) the presence of key features of the EAP already embedded in usual care (UC). METHODS: Five trained GPs were videotaped while treating a standardized patient with EAP. Additionally, five GPs administering UC were videotaped. Two blinded observers evaluated the videotapes in relation to key features of the EAP which were scored on the EAP checklist. RESULTS: The mean total score on the EAP checklist was 4.7 (SD = 2.9) for the UC group and 7.1 (SD = 2.1) for the EAP group. Neither group reached a score higher than 8, which was considered to reflect an acceptable number of key EAP features. CONCLUSION: Our comparison of the presence of key features of EAP shows that the UC and EAP groups differed less than was expected. GPs in the UC group performed above expectation, with a mean total score of 4.7. Moreover, the low number of key features present in the EAP group may very well have led to a reduced effectiveness of the EAP. The results of this study can be used to optimize the training of GPs using the EAP

International consensus recommendations for management of new onset refractory status epilepticus including febrile infection-related epilepsy syndrome: Statements and supporting evidence

Objective: This study was undertaken to develop consensus-based recommendations for the management of adult and pediatric patients with new onset refractory status epilepticus (NORSE)/febrile infection-related epilepsy syndrome (FIRES) based on best evidence and experience. Methods: The Delphi methodology was followed. A facilitator group of nine experts was established, who defined the scope, users, and suggestions for recommendations. Following a review of the current literature, recommendation statements concerning diagnosis, treatment, and research directions were generated, which were then rated on a scale of 1 (strongly disagree) to 9 (strongly agree) by a panel of 48 experts in the field. Consensus that a statement was appropriate was reached if the median score was ≥7 and inappropriate if the median score was ≤3. The analysis of evidence was mapped to the results of each statement included in the Delphi survey. Results: Overall, 85 recommendation statements achieved consensus. The recommendations are divided into five sections: (1) disease characteristics; (2) diagnostic testing and sampling; (3) acute treatment; (4) treatment in the postacute phase; and (5) research, registries, and future directions in NORSE/FIRES. The detailed results and discussion of all 85 statements are outlined herein. A corresponding summary of findings and practical flowsheets are presented in a companion article. Significance: This detailed analysis offers insight into the supporting evidence and the current gaps in the literature that are associated with expert consensus statements related to NORSE/FIRES. The recommendations generated by this consensus can be used as a guide for the diagnosis, evaluation, and management of patients with NORSE/FIRES, and for planning of future research

The Hawthorne Effect: a randomised, controlled trial

Background: The 'Hawthorne Effect' may be an important factor affecting the generalisability of clinical research to routine practice, but has been little studied. Hawthorne Effects have been reported in previous clinical trials in dementia but to our knowledge, no attempt has been made to quantify them. Our aim was to compare minimal follow- up to intensive follow-up in participants in a placebo controlled trial of Ginkgo biloba for treating mild-moderate dementia.Methods: Participants in a dementia trial were randomised to intensive follow- up (with comprehensive assessment visits at baseline and two, four and six months post randomisation) or minimal follow-up (with an abbreviated assessment at baseline and a full assessment at six months). Our primary outcomes were cognitive functioning (ADAS-Cog) and participant and carer-rated quality of life (QOL-AD).Results: We recruited 176 participants, mainly through general practices. The main analysis was based on Intention to treat (ITT), with available data. In the ANCOVA model with baseline score as a co- variate, follow-up group had a significant effect on outcome at six months on the ADAS-Cog score (n = 140; mean difference = -2.018; 95% Cl -3.914, -0.121; p = 0.037 favouring the intensive follow-up group), and on participant- rated quality of life score (n = 142; mean difference = -1.382; 95% Cl -2.642, -0.122; p = 0.032 favouring minimal follow-up group). There was no significant difference on carer quality of life.Conclusion: We found that more intensive follow-up of individuals in a placebo-controlled clinical trial of Ginkgo biloba for treating mild-moderate dementia resulted in a better outcome than minimal follow-up, as measured by their cognitive functioning

International Consensus Recommendations for the Treatment of Pediatric NMDAR Antibody Encephalitis.

OBJECTIVE: To create an international consensus treatment recommendation for pediatric NMDA receptor antibody encephalitis (NMDARE). METHODS: After selection of a panel of 27 experts with representation from all continents, a 2-step Delphi method was adopted to develop consensus on relevant treatment regimens and statements, along with key definitions in pediatric NMDARE (disease severity, failure to improve, and relapse). Finally, an online face-to-face meeting was held to reach consensus (defined as ≥75% agreement). RESULTS: Corticosteroids are recommended in all children with NMDARE (pulsed IV preferred), with additional IV immunoglobulin or plasma exchange in severe patients. Prolonged first-line immunotherapy can be offered for up to 3-12 months (oral corticosteroids or monthly IV corticosteroids/immunoglobulin), dependent on disease severity. Second-line treatments are recommended for cases refractory to first-line therapies (rituximab preferred over cyclophosphamide) and should be considered about 2 weeks after first-line initiation. Further immunotherapies for refractory disease 1-3 months after second-line initiation include another second-line treatment (such as cyclophosphamide) and escalation to tocilizumab. Maintenance immune suppression beyond 6 months (such as rituximab redosing or mycophenolate mofetil) is generally not required, except for patients with a more severe course or prolonged impairments and hospitalization. For patients with relapsing disease, second-line and prolonged maintenance therapy should be considered. The treatment of NMDARE following herpes simplex encephalitis should be similar to idiopathic NMDARE. Broad guidance is provided for the total treatment duration (first line, second line, and maintenance), which is dictated by the severity and clinical course (i.e., median 3, 9 and 18 months in the best, average, and worst responders, respectively). Recommendations on the timing of oncologic searches are provided. CONCLUSION: These international consensus recommendations for the management of pediatric NMDARE aim to standardize the treatment and provide practical guidance for clinicians, rather than absolute rules. A similar recommendation could be applicable to adult patients

Supervising the Supervisors—Procedural Training and Supervision in Internal Medicine Residency

At teaching hospitals, bedside procedures (paracentesis, thoracentesis, lumbar puncture, arthrocentesis and central venous catheter insertion) are performed by junior residents and supervised by senior peers. Residents’ perceptions about supervision or how often peer supervision produces unsafe clinical situations are unknown. To examine the experience and practice patterns of residents performing bedside procedures. Cross-sectional e-mail survey of 653 internal medicine (IM) residents at seven California teaching hospitals. Surveys asked questions in three areas: (1) resident experience performing procedures: numbers of procedures performed and whether they received other (e.g., simulator) training; (2) resident comfort performing and supervising procedures; (3) resident reports of their current level of supervision doing procedures, experience with complications as well as perceptions of factors that may have contributed to complications. Three hundred sixty-seven (56%) of the residents responded. Most PGY1 residents had performed fewer than five of any of the procedures, but most PGY-3 residents had performed at least ten by the end of their training. Resident comfort for each procedure increased with the number of procedures performed (p < 0.001). Although residents reported that peer supervision happened often, they also reported high rates of supervising a procedure before feeling comfortable with proper technique. The majority of residents (64%) reported at least one complication and did not feel supervision would have prevented complications, even though many reported complications represented technique- or preparation-related problems. Residents report low levels of comfort and experience with procedures, and frequently report supervising prior to feeling comfortable. Our findings suggest a need to examine best practices for procedural supervision of trainees