Modular 5'-UTR hexamers for context-independent tuning of protein expression in eukaryotes.

Functional characterization of regulatory DNA elements in broad genetic contexts is a prerequisite for forward engineering of biological systems. Translation initiation site (TIS) sequences are attractive to use for regulating gene activity and metabolic pathway fluxes because the genetic changes are minimal. However, limited knowledge is available on tuning gene outputs by varying TISs in different genetic and environmental contexts. Here, we created TIS hexamer libraries in baker's yeast Saccharomyces cerevisiae directly 5' end of a reporter gene in various promoter contexts and measured gene activity distributions for each library. Next, selected TIS sequences, resulted in almost 10-fold changes in reporter outputs, were experimentally characterized in various environmental and genetic contexts in both yeast and mammalian cells. From our analyses, we observed strong linear correlations (R2 = 0.75-0.98) between all pairwise combinations of TIS order and gene activity. Finally, our analysis enabled the identification of a TIS with almost 50% stronger output than a commonly used TIS for protein expression in mammalian cells, and selected TISs were also used to tune gene activities in yeast at a metabolic branch point in order to prototype fitness and carotenoid production landscapes. Taken together, the characterized TISs support reliable context-independent forward engineering of translation initiation in eukaryotes.

Voxel-Based Dosimetry of Iron Oxide Nanoparticle-Conjugated 177Lu-Labeled Folic Acid Using SPECT/CT Imaging of Mice.

Several radiolabeled folic acid conjugates have been developed for targeted imaging and therapy. However, the therapeutic concept with radiolabeled folate conjugates has not yet been applied to clinical applications owing to the high renal absorbed dose. The effectiveness of targeted radionuclide therapy (TRT) depends primarily on the absorbed dose rate and on the total absorbed dose delivered to the tumor and to normal tissue. Owing to various limitations associated with organ level dosimetry, voxel-based dosimetry has become essential for the assessment of a more  accurate absorbed dose during TRT. In this study, we synthesized iron oxide nanoparticle (IONP)-conjugated radiolabeled folate (177Lu-IONP-Folate) and performed voxel-based dosimetry using SPECT/CT images of normal mice through direct Geant4 application for emission tomography (GATE) Monte Carlo (MC) simulation. We also prepared 177Lu-Folate and 177Lu-IONPs for the comparison of absorbed doses with that of 177Lu-IONP-Folate. In addition, we calculated the mean absorbed dose at the organ-level using the medical internal radiation dose (MIRD) schema. The radioactivities of all three radiotracers were mainly accumulated in the liver and kidneys immediately after injection. For the kidneys, the voxel-based absorbed doses obtained with 177Lu-IONP-Folate, 177Lu-Folate, and 177Lu-IONPs were 1.01 ± 0.17, 2.46 ± 0.50, and 0.52 ± 0.08 Gy/MBq, respectively. The renal absorbed dose decreased significantly (∼half) when 177Lu-IONP-Folate was used compared with when the 177Lu-Folate only was used. The mean absorbed dose values obtained at organ-level using the MIRD schema were comparable to voxel-based absorbed doses estimated with GATE MC. The voxel-based absorbed dose values obtained in this study of individualized activity show that the renal absorbed dose could be reduced to almost half with 177Lu-IONP-Folate. Therefore, 177Lu-IONP-Folate could be clinically applicable in the TRT of folate receptor-positive cancers in a personalized manner when using the voxel-based dosimetry method.

Histomorphological factors in the risk prediction of lymph node metastasis in papillary thyroid carcinoma.

AIMS: Few clinicopathological parameters have been identified as independent predictive factors for lymph node metastasis. This study evaluated the predictive ability of three histological characteristics of PTC in lymph node metastases: hobnail features, loss of cohesiveness/polarity (LOCP) and micropapillary structures. METHODS AND RESULTS: Tissue specimens from 153 patients with histologically confirmed PTC including 112 cases of papillary thyroid microcarcinoma (PTMC) were enrolled in this study. Three histological characteristics (hobnail features, LOCP and micropapillary structures) and several clinicopathological parameters were evaluated for their value in predicting lymph node metastasis. Hobnail features, LOCP and micropapillary structures were each significantly associated with and found to be independent predictive factors for lymph node metastasis (P < 0.05). These three histological characteristics were closely correlated with one another (P < 0.001). Six of the seven possible combinations of these three histological characteristics were independently correlated with lymph node metastasis (P < 0.05). Among these combinations, the coincidence of all three histological parameters represented the strongest independent predictive factor for lymph node metastasis (OR: 3.270, P = 0.006). CONCLUSIONS: Our study demonstrates that hobnail features, LOCP and micropapillary structures, either alone or in combinations, represent strong independent predictive factors for lymph node metastasis in PTC.

Control of Tire Wear Particulate Matter through Tire Tread Prescription.

This study aims to analyze tire wear particulate matter (TWP) from tread rubber with different formulations and to compare the concentration of TWP with different wear devices. The TWP generated during the abrasion of truck and bus radial (TBR) tires were examined, and the effect of using different types of rubber and carbon black (CB) were investigated. When natural rubber (NR) was solely used as the tire tread rubber material, there was a higher concentration of 5-10 µm TWP. However, when the tread formulation consisted of NR mixed with butadiene rubber, the TWP concentration decreased. Changing the type of CB also reduced the amount of TWP in the 2.5 µm size range. The TWP concentration in the specimens increased with increasing speed and vertical load. The TWP generated during the abrasion tests using wear testers and tire simulators exhibited similar trends. These findings suggest that modifying tire tread formulations can effectively control the distribution and amount of TWP generation.

Chronic Dantrolene Treatment Does Not Affect Hypertension, but Attenuates Sympathetic Stimulation Enhanced Atrial Fibrillation Inducibility in SHR.

BACKGROUND: Ryanodine receptor (RyR) dysfunction in skeletal muscle (RyR1) leads to malignant hyperthermia, and in cardiac muscle (RyR2) triggers cardiac arrhythmias. We hypothesized that RyR dysfunction in vascular smooth muscle could increase vascular resistance and hypertension, and may contribute to increased atrial fibrillation (AF) in hypertension. Thus, stabilizing RyR function with chronic dantrolene treatment may attenuate hypertension and AF inducibility in spontaneously hypertensive rats (SHR). METHODS: Male SHR (16 weeks old) were randomized into vehicle- (n = 10) and dantrolene-treated (10 mg/kg/day, n = 10) groups for 4 weeks. Wistar Kyoto (WKY, n = 11) rats served as controls. Blood pressures (BP) were monitored before and during the 4-week treatment. After 4-week treatment, direct BP, echocardiography, and hemodynamics were recorded. AF inducibility tests were performed in vivo at baseline and repeated under sympathetic stimulation (SS). RESULTS: Compared with WKY, SHR had significantly higher BP throughout the experimental period. Dantrolene treatment had no effect on BP levels in SHR (final systolic BP 212 ± 9 mm Hg in vehicle group vs. 208 ± 16 mm Hg in dantrolene group, P > 0.05). AF inducibility was very low and not significantly different between 5-month-old WKY and SHR at baseline. However, under SS, AF inducibility and duration were significantly increased in SHR (20% in WKY vs. 60% in SHR-vehicle, P<0.05). Dantrolene treatment significantly attenuated AF inducibility under SS in SHR (60% in vehicle vs. 20% in dantrolene, P < 0.05). CONCLUSIONS: Stabilizing RyR with chronic dantrolene treatment does not affect hypertension development in SHR. SHR has increased vulnerability to AF induction under SS, which can be attenuated with dantrolene treatment.

Formation of buried superconducting Mo2N by nitrogen-ion-implantation.

Nitrogen ion implantation is a useful technique to put nitrogen ions into lattices. In this work, nitrogen ion implantation into epitaxial Mo films is performed to create a buried superconducting γ-Mo2N. Atomically flat epitaxial (110) Mo films are grown on (0001) Al2O3. By impinging nitrogen ions, where the beam energy is fixed to 20 keV, we observe (111) γ-Mo2N diffraction and the formation of a γ-Mo2N layer from X-ray reflectivity. Magnetization and transport measurements clearly support a superconducting layer in the implanted film. Our strategy shows that formation of a buried superconducting layer can be achieved through ion implantation and self-annealing.

The correlation between shape of the sigmoid notch of the distal radius and the risk of triangular fibrocartilage complex foveal tear.

AIMS: The aim of this study was to analyze the association between the shape of the distal radius sigmoid notch and triangular fibrocartilage complex (TFCC) foveal tear. METHODS: Between 2013 and 2018, patients were retrospectively recruited in two different groups. The patient group comprised individuals who underwent arthroscopic transosseous TFCC foveal repair for foveal tear of the wrist. The control group comprised individuals presenting with various diseases around wrist not affecting the TFCC. The study recruited 176 patients (58 patients, 118 controls). The sigmoid notch shape was classified into four types (flat-face, C-, S-, and ski-slope types) and three radiological parameters related to the sigmoid notch (namely, the radius curvature, depth, and version angle) were measured. The association of radiological parameters and sigmoid notch types with the TFCC foveal tear was investigated in univariate and multivariate analyses. Receiver operating characteristic curves were used to estimate a cut-off for any statistically significant variables. RESULTS: Univariate analysis showed that the flat-face type was more prevalent in the patients than in the control group (43% vs 21%; p = 0.002), while the C-type was lower in the patients than in the control group (3% vs 17%; p = 0.011). The depth and version angle of sigmoid notch showed a negative association with the TFCC foveal tear in the multivariate analysis (depth: odds ratio (OR) 0.380; p = 0.037; version angle: OR 0.896; p = 0.033). Estimated cut-off values were 1.34 mm for the depth (area under the curve (AUC) = 0.725) and 10.45° for the version angle (AUC = 0.726). CONCLUSION: The proportion of flat-face sigmoid notch type was greater in the patient group than in the control group. The depth and version angle of sigmoid notch were negatively associated with TFCC foveal injury. Cite this article: Bone Joint J 2020;102-B(6):749-754.

MRI-based assessment of proximal femur strength compared to mechanical testing.

Half of the women who sustain a hip fracture would not qualify for osteoporosis treatment based on current DXA-estimated bone mineral density criteria. Therefore, a better approach is needed to determine if an individual is at risk of hip fracture from a fall. The objective of this study was to determine the association between radiation-free MRI-derived bone strength and strain simulations compared to results from direct mechanical testing of cadaveric femora. Imaging was conducted on a 3-Tesla MRI scanner using two sequences: one balanced steady-state free precession sequence with 300 µm isotropic voxel size and one spoiled gradient echo with anisotropic voxel size of 234 × 234 × 1500 µm. Femora were dissected free of soft-tissue and 4350-ohm strain-gauges were securely applied to surfaces at the femoral shaft, inferior neck, greater trochanter, and superior neck. Cadavers were mechanically tested with a hydraulic universal test frame to simulate loading in a sideways fall orientation. Sideways fall forces were simulated on MRI-based finite element meshes and bone stiffness, failure force, and force for plastic deformation were computed. Simulated bone strength metrics from the 300 µm isotropic sequence showed strong agreement with experimentally obtained values of bone strength, with stiffness (r = 0.88, p = 0.0002), plastic deformation point (r = 0.89, p < 0.0001), and failure force (r = 0.92, p < 0.0001). The anisotropic sequence showed similar trends for stiffness, plastic deformation point, and failure force (r = 0.68, 0.70, 0.84; p = 0.02, 0.01, 0.0006, respectively). Surface strain-gauge measurements showed moderate to strong agreement with simulated magnitude strain values at the greater trochanter, superior neck, and inferior neck (r = -0.97, -0.86, 0.80; p ≤0.0001, 0.003, 0.03, respectively). The findings from this study support the use of MRI-based FE analysis of the hip to reliably predict the mechanical competence of the human femur in clinical settings.

Diagnosis and Monitoring of Osteoporosis With 18F-Sodium Fluoride PET: An Unavoidable Path for the Foreseeable Future.

The prevalence of metabolic bone diseases particularly osteoporosis and its precursor, osteopenia, continue to grow as serious global health issues today. On a worldwide perspective, 200million people suffer from osteoporosis and in 2005, over 2million fracture incidents were estimated due to osteoporosis in the United States. Currently, osteoporosis and other metabolic bone diseases are evaluated primarily through dual energy X-ray absorptiometry, and rarely by bone biopsy with tetracycline labeling or Technetium-99m (99mTc) based bone scintigraphy. Deficiencies in these methods have prompted the use of more precise methods of assessment. This review highlights the use of 18F-sodium fluoride (NaF) with PET (NaF-PET), NaF-PET/CT, or NaF-PET/MRI in the evaluation of osteoporosis and osteopenia in the lumbar spine and hip. This imaging modality provides a molecular perspective with respect to the underlying metabolic alterations that lead to osseous disorders by measuring bone turnover through standardized uptake values. Its sensitivity and ability to examine the entire skeletal system make it a more superior imaging modality compared to standard structural imaging techniques. Further research is needed to determine its accuracy in reflecting the efficacy of therapeutic interventions in metabolic bone diseases.

A comprehensive listing of bioactivation pathways of organic functional groups.

The occurrence of idiosyncratic adverse drug reactions during late clinical trials or after a drug has been released can lead to a severe restriction in its use and even in its withdrawal. Metabolic activation of relatively inert functional groups to reactive electrophilic intermediates is considered to be an obligatory event in the etiology of many drug-induced adverse reactions. Therefore, a thorough examination of the biochemical reactivity of functional groups/structural motifs in all new drug candidates is essential from a safety standpoint. A major theme attempted in this review is the comprehensive cataloging of all of the known bioactivation pathways of functional groups or structural motifs commonly utilized in drug design efforts. Potential strategies in the detection of reactive intermediates in biochemical systems are also discussed. The intention of this review is not to "black list" functional groups or to immediately discard compounds based on their potential to form reactive metabolites, but rather to serve as a resource describing the structural diversity of these functionalities as well as experimental approaches that could be taken to evaluate whether a "structural alert" in a new drug candidate undergoes bioactivation to reactive metabolites.

Comparative assessment of parametric neuroreceptor mapping approaches based on the simplified reference tissue model using [¹¹C]ABP688 PET.

In recent years, several linearized model approaches for fast and reliable parametric neuroreceptor mapping based on dynamic nuclear imaging have been developed from the simplified reference tissue model (SRTM) equation. All the methods share the basic SRTM assumptions, but use different schemes to alleviate the effect of noise in dynamic-image voxels. Thus, this study aimed to compare those approaches in terms of their performance in parametric image generation. We used the basis function method and MRTM2 (multilinear reference tissue model with two parameters), which require a division process to obtain the distribution volume ratio (DVR). In addition, a linear model with the DVR as a model parameter (multilinear SRTM) was used in two forms: one based on linear least squares and the other based on extension of total least squares (TLS). Assessment using simulated and actual dynamic [(11)C]ABP688 positron emission tomography data revealed their equivalence with the SRTM, except for different noise susceptibilities. In the DVR image production, the two multilinear SRTM approaches achieved better image quality and regional compatibility with the SRTM than the others, with slightly better performance in the TLS-based method.

An fMRI investigation of hand representation in paraplegic humans.

OBJECTIVE: Cortical reorganization can occur after deaf-ferentation due to loss of a limb, but the nature of the cortical reorganization after spinal cord injury (SCI) is still in debate. METHODS: Using a 1.5T MRI, we scanned paraplegic and noninjured participants during hand movement and palm stimulation, to determine whether longterm paraplegics would show different patterns of cortical activity from the noninjured participants. RESULTS: The SCI group showed stronger activation in areas posterior, rather than superior, to the areas activated by non-SCIs. Conversely, the non-SCIs showed stronger activation in more anterior areas. The signal at each individual's maximally significant voxel had a greater modulation for the SCI group than for the non-SCIs, in response to movement. CONCLUSIONS: In this study of sensory and motor representations within the same subjects, the authors show for the first time the increase in the BOLD fMRI signal modulation in SCI. The authors do not find evidence of expansion of the hand representation into nearby cortical areas, and they corroborate previous EEG studies indicating a posterior shift for hand motor representation after SCI, while showing that the sensory representation does not undergo a posterior shift of similar magnitude. The difference between the reorganization found here and the reorganization typically found following amputations suggests a rationale for the differences in neuropathic pain symptoms following a spinal cord injury or amputation.

P-glycoprotein limits the brain penetration of nonsedating but not sedating H1-antagonists.

The present study evaluates the impact of P-glycoprotein (P-gp) on plasma-brain disposition and transepithelial transport of sedating versus nonsedating H1-antagonists using multidrug-resistant (mdr) gene 1a and 1b (mdr1a/b) knockout (KO) mice and human MDR1-transfected Madin-Darby canine kidney (MDCK) cells. Three nonsedating (cetirizine, loratadine, and desloratadine) and three sedating (diphenhydramine, hydroxyzine, and triprolidine) H1-antagonists were tested. Each compound was administered to KO and wild-type (WT) mice intravenously at 5 mg/kg. Plasma and brain drug concentrations were determined by liquid chromatography-mass spectrometry analysis. Mean pharmacokinetic parameters (CL, V(ss), and t(1/2)) were obtained using WinNonlin. In addition, certirizine, desloratadine, diphenhydramine, and triprolidine (2 microM) were tested as substrates for MDR1 using MDR1-MDCK cells. The bidirectional apparent permeability was determined by measuring the amount of compound at the receiving side at 5 h. The brain-to-plasma area under the curve (AUC) ratio was 4-, 2-, and >14-fold higher in KO compared with WT mice for cetirizine, loratadine, and desloratadine, respectively. In contrast, the brain-to-plasma AUC ratio between KO and WT was comparable for hydroxyzine, diphenhydramine, and triprolidine. Likewise, the efflux ratio between basolateral to apical and apical to basolateral was 4.6- and 6.6-fold higher in MDR1-MDCK than the parental MDCK for certirizine and desloratadine, respectively, whereas it was approximately 1 for diphenhydramine and triprolidine. Our results demonstrate that sedating H1-antagonists hydroxyzine, diphenhydramine, and triprolidine are not P-gp substrates. In contrast, nonsedating H1-antagonists cetirizine, loratadine, and desloratadine are P-gp substrates. Affinity for P-gp at BBB may explain the lack of central nervous system side effects of modern H1-antagonists.

Influence of the apolipoprotein E type 4 allele on cerebral glucose metabolism in Alzheimer's disease patients.

It is still unclear whether apolipoprotein E epsilon 4 (APOE E4) influences the cerebral glucose metabolism abnormalities found in Alzheimer's disease (AD), although APOE E4 is a well-known risk factor for AD. [(18)F]Fluorodeoxyglucose PET was conducted in patients with very mild (n=17), mild (n=27), and moderate-to-severe (n=19) AD. The presence of APOE E4 was associated with greater reduction of glucose metabolism in the left inferior temporal region in very mild AD but neither in mild nor in moderate-to-severe AD. These findings favor the hypothesis that APOE E4 is related mainly to the development of AD, not to its progression.