Impact of total thyroidectomy on quality of life at 6 months: the prospective ThyrQoL multicentre trial.

OBJECTIVE: This study is to determine the impact of complications after total thyroidectomy on health-related quality of life (HR-QoL) and to identify significant predictive factors of HR-QoL changes. HR-QoL is usually impaired in patients with thyroid diseases compared to the general population. Thyroidectomy is largely performed in the case of benign thyroid benign and can be associated with long-term complications (vocal cord palsy, hypoparathyroidism). DESIGN: The prospective ThyrQoL multicenter trial (NCT02167529) included 800 patients who underwent total thyroidectomy for benign or malignant non-extensive disease in seven French referral hospitals between 2014 and 2016. METHODS: HR-QoL was assessed using the MOS 36-item short form health survey (SF-36) self-questionnaire with a 6-month follow-up. RESULTS: We observed a significant improvement of HR-QoL 6 months after surgery (P < 0.0001). Postoperative complications were associated with a non-significant impairment of HR-QoL. In multivariable analysis, Graves' disease was associated with a significant improvement of HR-QoL (OR = 2.39 [1.49; 3.84]) and thyroid malignant disease with an impairment of HR-QoL (OR = 1.44 [0.99; 2.08]) after thyroidectomy. CONCLUSION: We observed a significant improvement of HR-QoL 6 months after total thyroid surgery for benign thyroid disease.

Roux-en-Y gastric bypass reduces plasma cholesterol in diet-induced obese mice by affecting trans-intestinal cholesterol excretion and intestinal cholesterol absorption.

OBJECTIVE: Bariatric surgery appears as the most efficient therapeutic alternative in morbidly obese patients. In addition to its efficiency to decrease body weight, it also improves metabolic complications associated to morbid obesity, including dyslipidemia. Although the cholesterol-lowering effect varies with the bariatric procedures, the underlying molecular mechanisms remain poorly defined. This study aims to assess the consequence of both restrictive (sleeve gastrectomy; SG) and malabsorptive (Roux-en-Y gastric bypass; RYGB) procedures on cholesterol metabolism in mice. SUBJECTS: Ten-week-old C57BL6/J males were fed with a high-fat diet for 8-14 weeks before sleeve or RYGB surgery. RESULTS: SG has a modest and transient effect on plasma cholesterol levels, linked to a reduction in food intake. In contrast, modified RYGB led to a sustained ≈35% reduction in plasma cholesterol concentrations with a drastic increase in fecal cholesterol output. Mechanistically, RYGB exerts a synergystic effect on cholesterol metabolism by inducing the trans-intestinal cholesterol efflux and reducing the intestinal cholesterol absorption. CONCLUSIONS: In mice, RYGB, but not sleeve, strongly favors plasma cholesterol elimination by concomitantly increasing trans-intestinal cholesterol excretion and by decreasing intestinal cholesterol absorption. Our models open new perspective for deciphering the hypocholesterolemic effects of bariatric procedures.

Wound closure with Karydakis flap is decreasing the perioperative costs after pilonidal sinus excision as compared to lay-open approach.

PURPOSE: To assess the impact of a simple flap closing procedure by Karydakis flap (KF) after pilonidal sinus excision on the costs and healing time as compared to routine lay-open technique. METHOD: Out of 44 consecutive patients operated on for pilonidal excision (November 2013-March 2015), 17 had a Karydakis flap and 27 a lay-open procedure. For each patient, the length of stay, the operating time (OT), the time needed for complete healing and postoperative care resources were recorded. The global costs included OT, nursing care quantity, and modalities until complete scar healing. RESULTS: One reoperation in the lay-open group was necessary during the follow-up (8±5months). No recurrence occurred. Postoperative morbidity was similar in both groups. Results showed that KF global cost was inferior as compared to lay-open technique (941±178€ vs. 1601±399€; P=0.0001), KF healed faster (32±17 vs. 59±22days; P=0.0001), whereas OT was longer in KF group (16±7 vs. 25±4min; P=0.001). CONCLUSION: KF allows a faster healing time and a 41% lower cost than lay-open technique. Preferential use of KF rather than lay-open procedure could allow a significant health cost saving.

Therapeutic expression of hairpins targeting apolipoprotein B100 induces phenotypic and transcriptome changes in murine liver.

Constitutive expression of short hairpin RNAs (shRNAs) may cause cellular toxicity in vivo and using microRNA (miRNA) scaffolds can circumvent this problem. Previously, we have shown that embedding small interfering RNA sequences targeting apolipoprotein B100 (ApoB) in shRNA (shApoB) or miRNA (miApoB) scaffolds resulted in differential processing and long-term efficacy in vivo. Here we show that adeno-associated virus (AAV)-shApoB- or AAV-miApoB-mediated ApoB knockdown induced differential liver morphology and transcriptome expression changes. Our analyses indicate that ApoB knockdown with both shApoB and miApoB resulted in alterations of genes involved in lipid metabolism. In addition, in AAV-shApoB-injected animals, genes involved in immune system activation or cell growth and death were affected, which was associated with increased hepatocyte proliferation. Subsequently, in AAV-miApoB-injected animals, changes of genes involved in oxidoreductase activity, oxidative phosphorylation and nucleic bases biosynthetic processes were observed. Our results demonstrate that long-term knockdown of ApoB in vivo by shApoB or miApoB induces several transcriptome changes in murine liver. The increased hepatocyte profileration by AAV-shRNA may have severe long-term effects indicating that AAV-mediated RNA interference therapy using artificial miRNA may be a safer approach for familial hypercholesterolemia therapy.

Occupational outcome after surgery in patients with a rotator cuff tear due to a work-related injury or occupational disease. A series of 262 cases.

UNLABELLED: The goals of this study were to establish the occupational outcome after surgery in patients with a rotator cuff tear from a work-related injury (WRI) or occupational disease (OD) and determine which factors and conditions affected return to work. HYPOTHESIS: return to work was possible for this type of patient. This questionnaire-based study comprised 262 shoulders in 254 patients with a WRI/OD who had surgery performed on their shoulder between 2000 and 2005. The average age was 50.5 ± 6.4 years. The following variables were analysed: employment status (private sector, self-employed, government employee), type of work (non-manual, manual, heavy manual labour), nature of tendon injury and surgical technique (open, mini-open and arthroscopy). Return to work occurred in 59.5% of the cases. Factors that prevented return to work (40.4% of the cases) included retirement (14.1%), an unrelated medical condition (10.3%), and the outcome of the operated shoulder (16.0%). Age had an impact on return to work (P<5 × 10(-4)). The type of work and nature of tendon injury did not affect return to work, but did affect time away from work. Employment status and surgical technique had an effect on return to work, but not on time away from work. Age was a decisive factor for return to work. Retirement seemed to be the most common choice starting at 55 years of age. Arthroscopy seemed to have reduced the impact of the WRI on the results, particularly on the time away from work. A preoperative evaluation of the patient's probability of returning to work should be done based on occupational and injury features. There may be a longer delay in returning to work for certain profiles of work (manual labour) and tendon injury. Patient management can be improved by knowing the factors and conditions that influence return to work. LEVEL OF EVIDENCE: Level IV - Retrospective study.

Upgrade of the CATS sample changer on FIP-BM30A at the ESRF: towards a commercialized standard.

An upgraded version of the sample changer ;CATS' (Cryogenic Automated Transfer System) that was developed on the FIP-BM30A beamline at the ESRF is presented. At present, CATS is installed at SLS (three systems), BESSY (one system), DLS (two systems) and APS (four systems for the LSCAT beamline). It consists mainly of an automated Dewar with an assortment of specific grippers designed to obtain a fast and reliable mounting/dismounting rate without jeopardizing the flexibility of the system. The upgraded system has the ability to manage any sample standard stored in any kind of puck.

FIP: a highly automated beamline for multiwavelength anomalous diffraction experiments.

FIP is a French Collaborating Research Group (CRG) beamline at the European Synchrotron Radiation Facility (ESRF) dedicated exclusively to crystallography of biological macromolecules, with a special emphasis on multiwavelength anomalous diffraction data collection in the 0.7-1.81 A wavelength range. The optics, consisting of long cylindrical grazing-angle mirrors associated with a cryocooled double-crystal monochromator, delivers an optimal beam in the corresponding energy range. The high level of automation, which includes automated crystal centring, automated data-collection management and data processing, makes the use of this beamline very easy. This is illustrated by the large number of challenging structures that have been solved since 1999.

Comparison of the DNA binding characteristics of the related zinc finger proteins WT1 and EGR1.

The interactions of the related zinc finger proteins WT1 and EGR1 with DNA have been investigated using a quantitative binding assay. A recombinant peptide containing the four zinc fingers of WT1 binds to the dodecamer DNA sequence GCG-TGG-GCG-TGT with an apparent dissociation constant (Kd) of (1.14 +/- 0.09) x 10(-9) M under conditions of 0.1 M KCl, pH 7.5, at 22 degrees C. Under the same conditions, a recombinant peptide containing the three zinc fingers of EGR1 binds to the dodecamer sequence, the first nine bases comprising the EGR consensus binding site, with an apparent Kd of (3.55 +/- 0.24) x 10(-9) M. The nature of the equilibrium binding of each peptide to DNA was investigated as a function of temperature, pH, monovalent salt concentration, and divalent salt concentration. The interaction of WT1 with DNA is an entropy-driven process, while the formation of the EGR1-DNA complex is favored by enthalpy and entropy. The DNA binding activities of both proteins have broad pH optima centered at pH 8.0. The binding of both proteins to DNA shows similar sensitivity to ionic strength, with approximately 7.7 +/- 0.8 ion pairs formed in the EGR1-DNA complex and 9.2 +/- 1.8 ion pairs formed in the WT1-DNA complex. Results of measuring the effects of point mutations in the DNA binding site on the affinity of WT1 and EGR1 indicates a significant difference in the optimal binding sites: for EGR1, the highest affinity binding site has the sequence GNG-(T/G)GG-G(T/C)G, while for WT1 the highest affinity binding site has the sequence G(T/C)G-(T/G)GG-GAG-(T/C)G(T/C).

Effects of Denys-Drash syndrome point mutations on the DNA binding activity of the Wilms' tumor suppressor protein WT1.

A number of point mutations in the zinc finger domain of the Wilms' tumor suppressor protein WT1 have been isolated from the DNA of patients with Denys-Drash syndrome, an association of Wilms' tumor, nephropathy, and genital anomalies. To date, five different mutations that alter amino acids predicted to interact specifically with nucleotides in the target DNA sequence have been described. Two of these mutations are located in zinc finger 2 (R366H, R366C), and three are located in finger 3 (R394W, D396G, D396N). These five Denys-Drash mutations were introduced into WT1-ZFP, a recombinant polypeptide containing the zinc finger domain of WT1, and the effects of these mutations on DNA sequence specificity were determined using a selection, amplification, and binding (SAAB) assay. The SAAB assay was carried out using two different DNA templates, one with a randomized finger 2 subsite (GCG TGG NNN TGT) and one with a randomized finger 3 subsite (GCG NNN GCG TGT). A comparison of the DNA sequences selected by WT1-ZFP and by Denys-Drash mutants suggests that the point mutations reduce the sequence selectivity of the zinc finger protein. With the exception of the R394W mutant, the other Denys-Drash mutations selected one alternative sequence in addition to the wild-type DNA subsite sequence. The binding affinities of these proteins for their selected sequences were determined using a quantitative nitrocellulose filter binding assay. These results revealed that the wild-type WT1 binds with slightly higher affinity to sequences with GAG in the finger 2 subsite than sequences with the EGR-1 consensus GCG finger 2 subsite. With the exception of R394W, which appears to lack specific DNA binding activity, the Denys-Drash mutants bound to selected DNAs with 1.4-14-fold lower affinities than the wild-type WT1-ZFP. These results suggest that the clinical phenotype of Denys-Drash syndrome can be associated with a modest reduction in the DNA binding affinity of WT1.

Seryl-tRNA synthetase from Escherichia coli: functional evidence for cross-dimer tRNA binding during aminoacylation.

Escherichia coli seryl-tRNA synthetase (SerRS) is a homo-dimeric class II aminoacyl-tRNA synthetase. Each subunit is composed of two distinct domains: the N-terminal domain is a 60 A long, arm-like coiled coil structure built up of two antiparallel alpha-helices, whereas the C-terminal domain, the catalytic core, is an alpha-beta structure overlying a seven-stranded antiparallel beta-sheet. Deletion of the arm-like domain (SerRS delta 35-97) does not affect the amino acid activation step of the reaction, but reduces aminoacylation activity by more than three orders of magnitude. In the present study, it was shown that the formation of heterodimers from two aminoacylation defective homodimers, the N-terminal deletion and an active site mutant (SerRS E355Q), restored charging activity. The aminoacylation activity in a mixture containing the heterodimers was compared to that of solutions containing the same concentrations of homodimer. The activity of the mixture was eight times higher than the activities of the homodimer solutions, and reached 50% of the theoretical value that would be expected if 50% of the mixture was in the heterodimer form and assuming that a heterodimer contains only one active site. These results are in full agreement with the structural analysis of E. coli SerRS complexed with its cognate tRNA and provide functional evidence for the cross-dimer binding of tRNA in solution.

Seryl-tRNA synthetase from Escherichia coli: implication of its N-terminal domain in aminoacylation activity and specificity.

Escherichia coli seryl-tRNA synthetase (SerRS) a dimeric class II aminoacyl-tRNA synthetase with two structural domains charges specifically the five iso-acceptor tRNA(ser) as well as the tRNA(sec) (selC product) of E. coli. The N-terminal domain is a 60 A long arm-like coiled coil structure built of 2 long antiparallel a-h helices, whereas the C-terminal domain is a alpha-beta structure. A deletion of the N-terminal arm of the enzyme does not affect the amino acid activation step of the reaction, but reduces dramatically amino-acylation activity. The Kcat/Km value for the mutant enzyme is reduced by more than 4 orders of magnitude, with a nearly 30 fold increased Km value for tRNA(ser). An only slightly truncated mutant form (16 amino acids of the tip of the arm replaced by a glycine) has an intermediate aminoacylation activity. Both mutant synthetases have lost their specificity for tRNA(ser) and charge also non-cognate type 1 tRNA(s). Our results support the hypothesis that class II synthetases have evolved from an ancestral catalytic core enzyme by adding non-catalytic N-terminal or C-terminal tRNA binding (specificity) domains which act as determinants for cognate and anti-determinants for non-cognate tRNAs.

Crystallization of the seryl-tRNA synthetase:tRNAS(ser) complex of Escherichia coli.

Crystals of the complex between seryl-tRNA synthetase and tRNA(2ser) from Escherichia coli have been obtained from ammonium sulphate solutions. The crystals are of the 1:2 enzyme:tRNA complex, belong to the space group C222(1), have cell dimensions of a = 128.9 A, b = 164.9 A, c = 127.3 A and diffract anisotropically from 3.5 to 4.5 A. An X-ray diffraction data set to 4 A has been collected. The combination of molecular replacement using the refined structure of the catalytic domain of the native enzyme, data from a heavy atom derivative and solvent flattening was used to produce a map at 4 A resolution. This shows that a tRNA molecule binds across the dimer, the anticodon stem and loop do not contact the protein and the helical arm of the enzyme contacts the T psi C loop and the long extra arm of the tRNA.

In vivo overexpression and purification of Escherichia coli tRNA(ser).

DNA fragments corresponding to the sequences of Escherichia coli tRNA(2ser) and amber suppressor tRNA(ser), were synthesized from overlapping oligonucleotides. These were interposed between a strong promotor and a synthetic transcriptional terminator to ensure the production of a transcript of the correct size. The genes of promotor, fragment and terminator were cloned into a conditional runaway replication plasmid. At temperatures below 37 degrees C this vector has a low copy number but, following a temperature shift to 42 degrees C, the copy number is no longer regulated. Using these constructs an overexpression of tRNA(ser) of about 20 times the level of the wild-type pool could be obtained (corresponding e.g. to 200 times the expression tRNA(2ser)). From these systems 10 mg quantities of tRNA(ser)s could be isolated with a serine acceptance of 1,100 pmol/A280 unit.

[Use of photogrammetry for taking body contours in radiotherapy]. / Emploi de la photogrammétrie pour la prise des contours corporels en radiothérapie

A photogrammetric method for taking outline of patients in radiotherapy is described in details. Using only one couple of photographs, this fast and very precise process may restitute a great deal of transversal or longitudinal cross-sections, even after a long time because photographic stocking of information. An automatic numeric lecture of the outline may directly enter a computer for dosimetry.