The [Lys(-2)-Arg(-1)-des(17-21)]-endothelin-1 peptide retains the specific Arg(-1)-Asp8 salt bridge but reveals discrepancies between NMR data and molecular dynamics simulations

The [des(17-21)]-endothelin-1 (CSH-ET) and [Lys(-)(2)-Arg(-)(1)-des(17-21)]-endothelin-1 (KR-CSH-ET) peptides, designed by removing the five-residue hydrophobic tail from the endothelin-1 (ET-1) and [Lys(-)(2)-Arg(-)(1)]-endothelin-1 (KR-ET-1) peptides, respectively, were synthesized. Previous studies on KR-ET-1 showed that, in contrast to ET-1, this engineered compound displays a pH-dependent conformational change related to the formation of a stabilizing salt bridge between the Arg(-)(1) and Asp(8) side chains. CD and NMR spectra indicate that CSH-ET and KR-CSH-ET display conformational behavior similar to those of ET-1 and KR-ET-1, respectively. The short salt bridge-stabilized KR-CSH-ET peptide therefore appears to be an attractive elementary scaffold for drug design. The solution structure of the salt-bridged form of KR-CSH-ET was determined by NMR at pH 4.5 and is very similar to the corresponding form of the parent KR-ET-1 peptide. Molecular dynamics simulations of the salt-bridged form of KR-CSH-ET were performed using both the GB/SA implicit solvation scheme or an explicit solvation and the particle-mesh Ewald method for long-range electrostatic calculation. Unexpectedly, the Arg(-)(1)-Asp(8) salt bridge does not display in the simulation the stability that could be expected from the experimental data. The cooperative involvement of a cation-pi interaction in formation of the salt bridge has been hypothesized. Difficulties in accurately simulating cation-pi interactions might be responsible for the lack of stability in the simulation. At this time, however, no definitive explanation for the observed discrepancy between experiments and simulations is available, and further experimental studies appear to be necessary to fully understand in atomic detail the pH-dependent conformational change observed in the KR-ET-1 series.     

In vivo behavior of muscle fascicles and tendinous tissues in human tibialis anterior muscle during twitch contraction

In this study we investigated the time course of length and velocity of muscle fascicles and tendinous tissues (TT) during isometric twitch contraction, and examined how their interaction relates to the time course of external torque and muscle fascicle force generation. From seven males, supra-maximal twitch contractions (singlet) of the tibialis anterior muscle were induced at 30 degrees , 10 degrees and -10 degrees plantar flexed positions. The length and velocity of fascicles and TT were determined from a series of their transverse ultrasound images. The maximal external torque appeared when the shortening velocity of fascicles was zero. The fascicle and TT length, and external torque showed a 10-30 ms delay of each onset, with a significant difference in half relaxation times at -10 degrees . The time course of TT elongation, and fascicle and tendinous velocities did not differ between joint angles. Curvilinear length-force properties, whose slope of quasi-linear part was ranged from -15.0 to -5.9 N/mm for fascicles and 5.4 to 14.3N/mm for TT, and a loop-like pattern of velocity-force properties, in which the mean power was ranged from 0.14 to 0.80 W for fascicles, and 0.14 to 0.81 W for TT were also observed. These results were attributed to the muscle-tendon interaction, depending on the slack and non-linearity of length-force relationship of compliant TT. We conclude that the mechanical interaction between fascicles and TT, are significant determinants of twitch force and time characteristics.     

Synthesis,SAR study,and biological evaluation of novel 2,3-dihydro-1H-imidazo[1,2-a]benzimidazole derivatives as phosphodiesterase 10A inhibitors

Phosphodiesterase 10A (PDE10A) inhibitors were designed and synthesized based on the dihydro-imidazobenzimidazole scaffold. Compound 5a showed moderate inhibitory activity and good permeability, but unfavorable high P-glycoprotein (P-gp) liability for brain penetration. We performed an optimization study to improve both the P-gp efflux ratio and PDE10A inhibitory activity. As a result, 6d was identified with improved P-gp liability and high PDE10A inhibitory activity. Compound 6d also showed satisfactory brain penetration, suppressed phencyclidine-induced hyperlocomotion and improved MK-801-induced working memory deficit.     

Adverse Events during Bowel Preparation and Colonoscopy in Patients with Acute Lower Gastrointestinal Bleeding Compared with Elective Non-Gastrointestinal Bleeding

Background

There are limited data on the safety of colonoscopy in patients with lower gastrointestinal bleeding (LGIB). We examined the various adverse events associated with colonoscopy in acute LGIB compared with non-GIB patients.

Methods

Emergency hospitalized LGIB patients (n = 161) and age- and gender-matched non-GIB controls (n = 161) were selected. Primary outcomes were any adverse events during preparation and colonoscopy procedure. Secondary outcomes were five bowel preparation-related adverse events–hypotension, systolic blood pressure <100 mmHg, volume overload, vomiting, aspiration pneumonia and loss of consciousness–and four colonoscopy-related adverse events–including hypotension, perforation, cerebrocardiovascular events and sepsis.

Results

During bowel preparation, 16 (9%) LGIB patients experienced an adverse event. None of the LGIB patients experienced volume overload, aspiration pneumonia or loss of consciousness; however, 12 (7%) had hypotension and 4 (2%) vomited. There were no significant differences in the five bowel preparation-related adverse events between LGIB and non-GIB patients. During colonoscopy, 25 (15%) LGIB patients experienced an adverse event. None LGIB patient had perforation or sepsis; however, 23 (14%) had hypotension and 2 (1%) experienced a cerebrocardiovascular event. There was no significant difference in the four colonoscopy-related adverse events between LGIB and non-GIB patients. In addition, no significant difference in any of the nine adverse events was found among subgroups: patients aged ≥65 years, those with comorbidities, and those with antithrombotic drug use.

Conclusions

Adverse events in bowel preparation and colonoscopy among acute LGIB patients were low. No significant difference was found in adverse events between LGIB and non-GIB patients. These adverse events were also low in elderly LGIB patients, as well as in those with co-morbidities and antithrombotic drug use, suggesting that colonoscopy performed during acute LGIB did not increase adverse events.     

The size index as a motor unit identifier in electromyography examined by numerical calculation.

A computer simulation was performed to investigate the size index as a motor unit identifier in electromyography. The size index calculated from the amplitude and area of the simulated motor unit action potential (MUP) was plotted against the distance between the needle electrode and current source to show how the index changes as a function of the distance. The index of the MUP also was plotted against the number of muscle fibers belonging to a single motor unit, the size of the motor unit territory, and the diameter of the muscle fibers in order to establish the major determinants of the index. The index was relatively constant for the distance less than 2 mm between the needle electrode and closest edge of the current source. It changed logarithmically with the number of muscle fibers and with the diameter of the fibers.     

Identification and molecular characterization of a chitinase from the hard tick Haemaphysalis longicornis

A cDNA encoding tick chitinase was cloned from a cDNA library of mRNA from Haemaphysalis longicornis eggs and designated as CHT1 cDNA. The CHT1 cDNA contains an open reading frame of 2790 bp that codes for 930 amino acid residues with a coding capacity of 104 kDa. The deduced amino acid sequence shows a 31% amino acid homology to Aedes aegypti chitinase and a multidomain structure containing one chitin binding peritrophin A domain and two glycosyl hydrolase family 18 chitin binding domains. The endogenous chitinase of H. longicornis was identified by a two-dimensional immunoblot analysis with mouse anti-rCHT1 serum and shown to have a molecular mass of 108 kDa with a pI of 5.0. A recombinant baculovirus AcMNPV.CHT1-expressed rCHT1 is glycosylated and able to degrade chitin. Chitin degradation was ablated by allosamidin in a dose-dependent manner. The optimal temperature and pH for activity of the purified chitinase were 45 degrees C and pH 5-7. The CHT1 cDNA has an ELR motif for chemokine-mediated angiogenesis and appears to be a chitinase of the chemokine family. Localization analysis using mouse anti-rCHT1 serum revealed that native chitinase is highly expressed in the epidermis and midgut of the tick. AcMNPV.CHT1 topically applied to H. longicornis ticks exhibited replication. This is the first report of insect baculovirus infection of ticks. The importance of AcMNPV.CHT1 as a novel bio-acaricide for tick control is discussed.     

Reliability and Validity of Quantifying Absolute Muscle Hardness Using Ultrasound Elastography

Muscle hardness is a mechanical property that represents transverse muscle stiffness. A quantitative method that uses ultrasound elastography for quantifying absolute human muscle hardness has been previously devised; however, its reliability and validity have not been completely verified. This study aimed to verify the reliability and validity of this quantitative method. The Young’s moduli of seven tissue-mimicking materials (in vitro; Young’s modulus range, 20–80 kPa; increments of 10 kPa) and the human medial gastrocnemius muscle (in vivo) were quantified using ultrasound elastography. On the basis of the strain/Young’s modulus ratio of two reference materials, one hard and one soft (Young’s moduli of 7 and 30 kPa, respectively), the Young’s moduli of the tissue-mimicking materials and medial gastrocnemius muscle were calculated. The intra- and inter-investigator reliability of the method was confirmed on the basis of acceptably low coefficient of variations (≤6.9%) and substantially high intraclass correlation coefficients (≥0.77) obtained from all measurements. The correlation coefficient between the Young’s moduli of the tissue-mimicking materials obtained using a mechanical method and ultrasound elastography was 0.996, which was equivalent to values previously obtained using magnetic resonance elastography. The Young’s moduli of the medial gastrocnemius muscle obtained using ultrasound elastography were within the range of values previously obtained using magnetic resonance elastography. The reliability and validity of the quantitative method for measuring absolute muscle hardness using ultrasound elastography were thus verified.     

Convergent synthesis of fluorescence-labeled probes of Annonaceous acetogenins and visualization of their cell distribution

The convergent synthesis of fluorescence-labeled solamin, an antitumor Annonaceous acetogenin, was accomplished by two asymmetric alkynylations of 2,5-diformyl tetrahydrofuran with an alkyne tagged with fluorescent groups and another alkyne with an α,β-unsaturated γ-lactone. Assay for the growth inhibitory activity against human cancer cell lines revealed that the probe with the fluorescent groups at the end of the hydrocarbon chain may have the same mode of action as natural acetogenins. The merged fluorescence of dansyl-labeled solamin and MitoTracker Red suggests that Annonaceous acetogenins localize in the mitochondria.