Characterization of Tryptophanase from Vibrio cholerae O1

AbstractTryptophanase (Trpase) encoded by the tnaA gene catalyzes the conversion of tryptophan to indole, which is an extracellular signaling molecule detected in various bacteria including Vibrio cholerae. Indole has been demonstrated to regulate biofilm formation, drug resistance, plasmid maintenance and spore formation of bacteria. In the present study, the tnaA gene from V. cholerae O1 (VcTrpase) was cloned and expressed in E. coli BL21(DE3) tn5:tnaA (a Trpase-deficient competent). VcTrpase was purified by Ni2+-NTA chromatography. The obtained VcTrpase had a molecular mass of approximately 49 kDa, a specific activity of 3 U/mg protein, and absorption peaks at 330 and 435nm. Using a site-directed mutagenesis technique, replacement of Arg419 by Val resulted in a VcTrpase completely devoid of activity. Thus, this site can be a target for drug design for controlling V. cholerae

Aortic valve calcification volumes and chronic brain infarctions in patients undergoing transcatheter aortic valve implantation

Chronic silent brain infarctions, detected as new white matter hyperintensities on magnetic resonance imaging (MRI) following transcatheter aortic valve implantation (TAVI), are associated with long-term cognitive deterioration. This is the first study to investigate to which extent the calcification volume of the native aortic valve (AV) measured with cardiac computed tomography angiography (CTA) predicts the increase in chronic white matter hyperintensity volume after TAVI. A total of 36 patients (79 ± 5 years, median EuroSCORE II 1.9%, Q1–Q3 1.5–3.4%) with severe AV stenosis underwent fluid attenuation inversion recovery (FLAIR) MRI < 24 h prior to TAVI and at 3 months follow-up for assessment of cerebral white matter hyperintensity volume (mL). Calcification volumes (mm3) of the AV, aortic arch, landing zone and left ventricle were measured on the CTA pre-TAVI. The largest calcification volumes were found in the AV (median 692 mm3) and aortic arch (median 633 mm3), with a large variation between patients (Q1–Q3 482–1297 mm3 and 213–1727 mm3, respectively). The white matter hyperintensity volume increased in 72% of the patients. In these patients the median volume increase was of 1.1 mL (Q1–Q3 0.3–4.6 mL), corresponding with a 27% increase from baseline (Q1–Q3 7–104%). The calcification volume in the AV predicted the increase of white matter hyperintensity volume (Δ%), with a 35% increase of white matter hyperintensity volume, per 100 mm3 of AV calcification volume (SE 8.5, p < 0.001). The calcification volumes in the aortic arch, landing zone and left ventricle were not associated with the increase in white matter hyperintensity volume. In 72% of the patients new chronic white matter hyperintensities developed 3 months after TAVI, with a median increase of 27%. A higher calcification volume in the AV was associated with a larger increase in the white matter hyperintensity volume. These findings show the potential for automated AV calcium screening as an imaging biomarker to predict chronic silent brain infarctions

A diffusion tensor-based method facilitating volumetric assessment of fiber orientations in skeletal muscle

Background The purpose of this study was to develop a DTI-based method to quantitatively assess fiber angles and changes therein in leg muscles in order to facilitate longitudinal studies on muscle fiber architectural adaptations in healthy subjects. Methods The upper legs of five volunteers were scanned twice on the same day. The right lower legs of five volunteers were scanned twice with the ankle in three positions, i.e. -15° dorsiflexion, 0° neutral position, and 30° plantarflexion. The MRI protocols consisted of a noise scan, a 3- point mDixon scan and a DTI scan. Fiber-angle color maps were generated for four muscles in the upper legs and two muscles in the lower leg. Voxel-wise fiber angles (θ) were calculated from the angle between the principal eigenvector of the diffusion tensor and a reference line defined between the origo and insertion points of each muscle. Bland-Altman analysis, intraclass correlation coefficient (ICC), coefficient of variation (CV%), minimal detectable change (MDC), standard error (SE) and Friedman test were used for assessing the feasibility of this method and in order to have an indication of the repeatability and the sensitivity. Results Bland-Altman analysis showed good repeatability (CV%<10 and 0.7≤ICC≤0.9) with exception of the Tibialis Anterior (TA) muscle in dorsiflexion position(CV%: 12.2) and the Semitendinosus (ST) muscle (left leg) (CV%: 11.4). The best repeatability metrics were found for the SOL muscle in neutral position (CV%: 2.6). Changes in average θ in TA and SOL with ankle positions were observed in accordance with expected agonist and antagonist functions of both muscles. For example, for the anterior left compartment the change in fiber angle Δθ with respect to the neutral position Δθ = -1.6° ± 0.8° and 2.2° ± 2.8° (p = 0.008), for dorsiflexion and plantarflexion, respectively. Conclusion Our method facilitates fast inspection and quantification of muscle fiber angles in the lower and upper leg muscles in rest and detection of changes in lower-leg muscle fiber angles with varying ankle angles

The repeatability of bilateral diffusion tensor imaging (DTI) in the upper leg muscles of healthy adults

Objectives: Assessment of the repeatability of diffusion parameter estimations in the upper leg muscles of healthy adults over the time course of 2 weeks, from a simultaneous bilateral upper leg DTI measurement. Methods: SE-EPI DTI datasets were acquired at 3 T in the upper legs of 15 active adults at a time interval of 2 weeks. ROIs were manually drawn for four quadriceps and three hamstring muscles of both legs. The following DTI parameters were analyzed: 1st, 2nd, and 3rd eigenvalue (λ1, λ2, and λ3), mean diffusivity (MD), and fractional anisotropy (FA). DTI parameters per muscle were calculated with and without intravoxel incoherent motion (IVIM) correction together with SNR levels per muscle. Bland-Altman plots and within-subject coefficient of variation (wsCV) were calculated. Left-right differences between muscles were assessed. Results: The Bland-Altman analysis showed good repeatability of all DTI parameters except FA for both the IVIM-corrected and standard data. wsCV values show that MD has the highest repeatability (4.5% IVIM; 5.6% standard), followed by λ2 (4.9% IVIM; 5.5% standard), λ1 (5.3% IVIM; 7.5% standard), and λ3 (5.7% IVIM; 5.7% standard). wsCV values of FA were 15.2% for the IVIM-corrected data and 13.9% for the standard analysis. The SNR (41.8 ± 16.0 right leg, 41.7 ± 17.1 left leg) and wsCV values were similar for the left and right leg and no left-right bias was detected. Conclusions: Repeatability was good for standard DTI data and slightly better for IVIM-corrected DTI data. Our protocol is suitable for DTI of the upper legs with overall good SNR. Key Points: • The presented DTI protocol is repeatable and therefore suitable for bilateral DT imaging of the upper legs. • Additional B1+calibrations improve SNR and repeatability. • Correcting for perfusion effects improves repeatability

A diffusion tensor-based method facilitating volumetric assessment of fiber orientations in skeletal muscle

Background The purpose of this study was to develop a DTI-based method to quantitatively assess fiber angles and changes therein in leg muscles in order to facilitate longitudinal studies on muscle fiber architectural adaptations in healthy subjects. Methods The upper legs of five volunteers were scanned twice on the same day. The right lower legs of five volunteers were scanned twice with the ankle in three positions, i.e. -15° dorsiflexion, 0° neutral position, and 30° plantarflexion. The MRI protocols consisted of a noise scan, a 3- point mDixon scan and a DTI scan. Fiber-angle color maps were generated for four muscles in the upper legs and two muscles in the lower leg. Voxel-wise fiber angles (θ) were calculated from the angle between the principal eigenvector of the diffusion tensor and a reference line defined between the origo and insertion points of each muscle. Bland-Altman analysis, intraclass correlation coefficient (ICC), coefficient of variation (CV%), minimal detectable change (MDC), standard error (SE) and Friedman test were used for assessing the feasibility of this method and in order to have an indication of the repeatability and the sensitivity. Results Bland-Altman analysis showed good repeatability (CV%<10 and 0.7≤ICC≤0.9) with exception of the Tibialis Anterior (TA) muscle in dorsiflexion position(CV%: 12.2) and the Semitendinosus (ST) muscle (left leg) (CV%: 11.4). The best repeatability metrics were found for the SOL muscle in neutral position (CV%: 2.6). Changes in average θ in TA and SOL with ankle positions were observed in accordance with expected agonist and antagonist functions of both muscles. For example, for the anterior left compartment the change in fiber angle Δθ with respect to the neutral position Δθ = -1.6° ± 0.8° and 2.2° ± 2.8° (p = 0.008), for dorsiflexion and plantarflexion, respectively. Conclusion Our method facilitates fast inspection and quantification of muscle fiber angles in the lower and upper leg muscles in rest and detection of changes in lower-leg muscle fiber angles with varying ankle angles

MR Spectroscopy-derived Proton Density Fat Fraction Is Superior to Controlled Attenuation Parameter for Detecting and Grading Hepatic Steatosis

Purpose: To prospectively compare the diagnostic accuracy of controlled attenuation parameter (CAP) obtained with transient elastography and proton density fat fraction (PDFF) obtained with proton magnetic resonance (MR) spectroscopy with results of liver biopsy in a cohort of adult patients suspected of having nonalcoholic fatty liver disease (NAFLD). Materials and Methods: The institutional review board approved this study. Informed consent was obtained from all patients. The authors evaluated 55 patients suspected of having NAFLD (40 men, 15 women). Patients had a median age of 52.3 years (interquartile range [IQR], 43.7-57.6 years) and a median body mass index of 27.8 kg/m(2) (IQR, 26.0-33.1 kg/m(2)). CAP and PDFF measurements were obtained on the same day, within 27 days of biopsy (IQR, 7-44 days). CAP and PDFF were compared between steatosis grades by using the Jonckheere-Terpstra test. Diagnostic accuracies of CAP and PDFF for grading steatosis were assessed with receiver operating characteristic (ROC) analysis. Within-weeks reproducibility (CAP and PDFF) and within-session repeatability were assessed with linear regression analyses, intraclass correlation coefficients, and coefficients of variation. Results: Steatosis grades at liver biopsy were distributed as follows: S0, five patients; S1, 24 patients; S2, 17 patients; and S3, nine patients. Both PDFF and CAP helped detect histologically proven steatosis (>= S1), but PDFF showed better diagnostic accuracy than CAP in terms of the area under the ROC curve (0.99 vs 0.77, respectively; P=.0334). PDFF, but not CAP, enabled the grading of steatosis (P <.0001). For within-weeks reproducibility, the intraclass correlation coefficient with PDFF was higher than that with CAP (0.95 vs 0.65, respectively; P=.0015); coefficients of variation were similar (19% vs 11%, P=.55). Within-session repeatability of CAP was good, with a coefficient of variation of 4.5%. Conclusion: MR spectroscopy-derived PDFF is superior to CAP in detecting and grading liver steatosis in human NAFL

Effect of two eccentric hamstring exercises on muscle architectural characteristics assessed with diffusion tensor MRI

Objectives: To evaluate the effect of a Nordic hamstring exercise or Diver hamstring exercise intervention on biceps femoris long head, semitendinosus and semimembranosus muscle's fascicle length and orientation through diffusion tensor imaging (DTI) with magnetic resonance imaging. Methods: In this three-arm, single-center, randomized controlled trial, injury-free male basketball players were randomly assigned to a Nordic, Diver hamstring exercise intervention or control group. The primary outcome was the DTI-derived fascicle length and orientation of muscles over 12 weeks. Results: Fifty-three participants were included for analysis (mean age 22 ± 7 years). Fascicle length in the semitendinosus over 12 weeks significantly increased in the Nordic-group (mean [M]: 20.8 mm, 95% confidence interval [95% CI]: 7.8 to 33.8) compared with the Control-group (M: 0.9 mm, 95% CI: −7.1 to 8.9), mean between-groups difference: 19.9 mm, 95% CI: 1.9 to 37.9, p = 0.026. Fascicle orientation in the biceps femoris long head over 12 weeks significantly decreased in the Diver-group (mean: -2.6°, 95% CI: −4.1 to −1.0) compared with the Control-group (mean: −0.2°, 95% CI: −1.4 to 1.0), mean between-groups difference: -2.4°, 95% CI: −4.7 to −0.1, p = 0.039. Conclusion: The Nordic hamstring exercise intervention did significantly increase the fascicle length of the semitendinosus and the Diver hamstring exercise intervention did significantly change the orientation of fascicles of the biceps femoris long head. As both exercises are complementary to each other, the combination is relevant for preventing hamstring injuries

Sexual Dimorphism in Hepatic, Adipose Tissue, and Peripheral Tissue Insulin Sensitivity in Obese Humans

Glucose and lipid metabolism differ between men and women, and women tend to have better whole-body or muscle insulin sensitivity. This may be explained, in part, by differences in sex hormones and adipose tissue distribution. Few studies have investigated gender differences in hepatic, adipose tissue, and whole-body insulin sensitivity between severely obese men and women. In this study, we aimed to determine the differences in glucose metabolism between severely obese men and women using tissue-specific measurements of insulin sensitivity. Insulin sensitivity was compared between age and body mass index (BMI)-matched obese men and women by a two-step euglycemic hyperinsulinemic clamp with infusion of [6,6-H-2(2)]glucose. Basal endogenous glucose production (EGP) and insulin sensitivity of the liver, adipose tissue, and peripheral tissues were assessed. Liver fat content was assessed by proton magnetic resonance spectroscopy in a subset of included subjects. We included 46 obese men and women (age, 48 +/- 2 vs. 46 +/- 2 years, p = 0.591; BMI, 41 +/- 1 vs. 41 +/- 1 kg/m(2), p = 0.832). There was no difference in basal EGP (14.4 +/- 1.0 vs. 15.3 +/- 0.5 mu mol.kg fat-free mass(-1) min(-1), p = 0.410), adipose tissue insulin sensitivity (insulin-mediated suppression of free fatty acids, 71.6 +/- 3.6 vs. 76.1 +/- 2.6%, p = 0.314), or peripheral insulin sensitivity (insulin-stimulated rate of disappearance of glucose, 26.2 +/- 2.1 vs. 22.7 +/- 1.7 mu mol.kg(-1).min(-1), p = 0.211). Obese men were characterized by lower hepatic insulin sensitivity (insulin-mediated suppression of EGP, 61.7 +/- 4.1 vs. 72.8 +/- 2.5% in men vs. women, respectively, p = 0.028). Finally, these observations could not be explained by differences in liver fat content (men vs. women, 16.5 +/- 3.1 vs. 16.0 +/- 2.5%, p = 0.913, n = 27). We conclude that obese men have lower hepatic, but comparable adipose tissue and peripheral tissue, insulin sensitivity compared to similarly obese women. Hepatic insulin resistance may contribute to the higher prevalence of diabetes in obese men. Further insight into the mechanisms underlying this gender difference may reveal novel targets for diabetes prevention and/or therap

Intestinal acetate and butyrate availability is associated with glucose metabolism in healthy individuals

Animal studies suggest that short-chain fatty acids acetate and butyrate are key players in the gut-brain axis and may affect insulin sensitivity. We investigated the association of intestinal acetate and butyrate availability (measured by butyryl-coenzyme A transferase (ButCoA) gene amount) with insulin sensitivity and secretion in healthy subjects from the HELIUS cohort study from the highest 15N = 30) and the lowest 15N = 30) intestinal ButCoA gene amount. The groups did not differ in insulin sensitivity or secretion. However, the high ButCoA group showed lower glucose and insulin peaks during the first 60 min after a meal and a higher nadir during the second 60 min (p < 0.01), suggesting delayed glucose adsorption from the small intestine. Our data suggest that chronically increased acetate and butyrate availability may improve glucose metabolism by delaying gastric emptying and intestinal adsorption. Future studies should further investigate the effect of acetate and butyrate interventions.Peer reviewe

Hyperthermia treatment planning for cervical cancer patients based on electrical conductivity tissue properties acquired in vivo with EPT at 3 T MRI

Introduction The reliability of hyperthermia treatment planning (HTP) is strongly dependent on the accuracy of the electric properties of each tissue. The values currently used are mostly based on ex vivo measurements. In this study, in vivo conductivity of human muscle, bladder content and cervical tumours, acquired with magnetic resonance-based electric properties tomography (MR-EPT), are exploited to investigate the effect on HTP for cervical cancer patients. Methods Temperature-based optimisation of five different patients was performed using literature-based conductivity values yielding certain antenna settings, which are then used to compute the temperature distribution of the patient models with EPT-based conductivity values. Furthermore, the effects of altered bladder and muscle conductivity were studied separately. Finally, the temperature-based optimisation was performed with patient models based on EPT conductivity values. Results The tumour temperatures for all EPT-based dielectric patient models were lower compared to the optimal tumour temperatures based on literature values. The largest deviation was observed for patient 1 with ΔT90 = -1.37 °C. A negative impact was also observed when the treatment was optimised based on the EPT values. For four patients ΔT90 was less than 0.6 °C; for one patient it was 1.5 °C. Conclusions Electric conductivity values acquired by EPT are higher than commonly used from literature. This difference has a substantial impact on cervical tumour temperatures achieved during hyperthermia. A higher conductivity in the bladder and in the muscle tissue surrounding the tumour leads to higher power dissipation in the bladder and muscle, and therefore to lower tumour temperatures