The value of prognostic clinical data in Bell’s palsy

SummaryElectroneurography (ENoG) and clinical staging are currently the methods of choice to indicate prognosis in Bell’s palsy, although ENoG is an electrophysiological test not universally available. Aim: Identify other options of prognostic evaluation based upon clinical aspects and minimal electrical stimulation test allowing prognostic measurement in almost any circumstances. Study design: historic cohort. Material and Method: Chart review of 1,521 cases of IPFP, analyzing the following clinical aspects: gender, age, paralyzed side, installation mode, previous symptoms, associated symptoms and minimal electrical stimulation test (Hilger test) and its statistical correlation to facial palsy evolution after 6 months. Results: Data indicated that patients above 60 years old had worse prognosis in comparison with patients under 30 years old. A progressive mode of paralysis installation, absence of previous symptoms, concomitant vertigo and response superior to 3.5 mA at minimum electrical stimulation test were also related to worse prognosis. On the other hand, the absence of concomitant symptoms, diminished tearing and sudden onset were related to better prognosis. Conclusion: Clinical factors and Hilger’s test can accurately indicate the prognosis in cases of Bell’s palsy when ENoG is not available

Increasing the chemical-shift dispersion of unstructured proteins with a covalent lanthanide shift reagent.

The study of intrinsically disordered proteins (IDPs) by NMR often suffers from highly overlapped resonances that prevent unambiguous chemical-shift assignments, and data analysis that relies on well-separated resonances. We present a covalent paramagnetic lanthanide-binding tag (LBT) for increasing the chemical-shift dispersion and facilitating the chemical-shift assignment of challenging, repeat-containing IDPs. Linkage of the DOTA-based LBT to a cysteine residue induces pseudo-contact shifts (PCS) for resonances more than 20 residues from the spin-labeling site. This leads to increased chemical-shift dispersion and decreased signal overlap, thereby greatly facilitating chemical-shift assignment. This approach is applicable to IDPs of varying sizes and complexity, and is particularly helpful for repeat-containing IDPs and low-complexity regions. This results in improved efficiency for IDP analysis and binding studies

Type-II NADH:quinone oxidoreductase from <em>Staphylococcus aureus </em>has two distinct binding sites and is rate limited by quinone reduction.

A prerequisite for any rational drug design strategy is understanding the mode of protein-ligand interaction. This motivated us to explore protein-substrate interaction in Type-II NADH:quinone oxidoreductase (NDH-2) from Staphylococcus aureus, a worldwide problem in clinical medicine due to its multiple drug resistant forms. NDHs-2 are involved in respiratory chains and recognized as suitable targets for novel antimicrobial therapies, as these are the only enzymes with NADH:quinone oxidoreductase activity expressed in many pathogenic organisms. We obtained crystal and solution structures of NDH-2 from S. aureus, showing that it is a dimer in solution. We report fast kinetic analyses of the protein and detected a charge-transfer complex formed between NAD(+) and the reduced flavin, which is dissociated by the quinone. We observed that the quinone reduction is the rate limiting step and also the only half-reaction affected by the presence of HQNO, an inhibitor. We analyzed protein-substrate interactions by fluorescence and STD-NMR spectroscopies, which indicate that NADH and the quinone bind to different sites. In summary, our combined results show the presence of distinct binding sites for the two substrates, identified quinone reduction as the rate limiting step and indicate the establishment of a NAD(+) -protein complex, which is released by the quinone

A Comparison Between Conventional Formalin and Novel PAXgene Tissue Fixation: the European Spidia Morphology Ring Trials Experience

Introduction and Aim: Molecular pathology is an emerging discipline requiring high quality tissue samples which allow simultaneous molecular and histopathological analysis. Within the European FP7 project (grant agreement no. 222916) \u201cStandardisation and improvement of generic pre-analytical tools and procedures for in-vitro diagnostics\u201d (SPIDIA) different morphology ring trials were implemented. The aim of these ring trials was to evaluate the quality of histomorphology and suitability of PAXgene-fixed and paraffin-embedded (PFPE) samples for routine diagnostics, with special emphasis on the tumor grading, in comparison to the current state-of-the-art technique for routine morphological diagnostics formaldehyde-fixed and paraffin-embedded (FFPE) samples in breast and colon cancer tissues. Methods: Each cancer tissue sample was divided into two mirrored samples for PFPE and FFPE. Haematoxylin and eosin and Periodic-Acid-Schiff stained sections were scanned and evaluated in a blinded, randomized ring trial by pathologists from Europe and the US using virtual microscopy. Each participant evaluated only one of the two virtual slides obtained from each case according to a randomized scheme implying the subdivision of both, cases and participants, in two groups. Links to the randomized virtual slides were distributed by email to the participants together with the study protocol, the electronic evaluation form and all instructions and timeline for the ring trial implementation. The reproducibility between fixation methods (PFPE vs. FFPE) was assessed by computing the modal category within each method for each case (Modal Scenario, MS). By starting from these values, the jackknifed estimate of the weighted kappa statistic (Kwj)1 was computed together with the relative 95% Confidence Interval (CI)2. In addition the inter-observer reproducibility was assessed by computing the Kwj and the kappa category-specific statistics (Kcs) and their weighted average (Cohen\u2019s kappa statistic, Kc) were estimated by jointly considering all participant\u2019s data1,3. Results: Tissue samples of 18 breast and 15 colon cancer cases were evaluated by 23 and 20 pathologists, respectively. By using the MS, a high and quite satisfactory level of reproducibility between PFPE and FFPE samples in evaluating the histological grading score was observed in breast (kwj=0.92, 95% CI: 0.76-1.00) and colon (kwj=0.73, 95% CI: 0.41-0.94) cancer, respectively. Moreover the inter-observer reproducibility for grading was not completely satisfactory in both ring trials, with G2 as the most critical category, but showed a similar performance level in each of the two considered groups of pathologists. Conclusions: Our findings suggest that the methodological approach we developed is suitable for evaluating the reproducibility between alternative fixation methods in the ring trial setting implemented here and that histomorphology is excellently preserved in PFPE tissues. References: 1. Fleiss JL. 2nd Ed. New York: Wiley and Sons 1981. 2. Corletto V, Verderio P, Giardini R et al. Anal Cell Pathol. 1998;16:83-93. 3. Holman CD. Am J Epidemiol. 1984;120:154-60. 4. Landis JR, Koch GG. Biometrics. 1977;33:159-74