A combined testing protocol approach for mutagenicity testing.

The antischistosomal agent, hycanthone methanesulfonate (HMS), was employed to illustrate the utility of carrying out several mutagenicity tests in a single concurrent animal experiment. Several commonly used procedures that were successfully integrated into a multiple testing protocol included (1) metaphase analysis in bone marrow, (2) micronucleus test in bone marrow, (3) analysis of the urine for mutagenic constituents, and (4) the host-mediated assay using Salmonella typhimurium. In addition to these animal studies, in vitro mutagenicity testing with and without activation was carried out using S. typhimurium. HMS produced positive, dose--response effects in in vitro tests, metaphase analysis, micronucleus test, and urine analysis, but not in the host-mediated assay. The results of these integrated techniques suggest that such a protocol may be a benefit to those concerned with mutagenicity testing of chemicals.     

Optimizing the exercise protocol for cardiopulmonary assessment

Twelve normal men performed 1-min incremental exercise tests to exhaustion in approximately 10 min on both treadmill and cycle ergometer. The maximal O2 uptake (VO2 max) and anaerobic threshold (AT) were higher (6 and 13%, respectively) on the treadmill than the cycle; the AT was reached at about 50% of VO2 max on both ergometers. Maximal CO2 output, heart rate, and O2 pulse were also slightly, but significantly higher on the treadmill. Maximal ventilation, gas exchange ratio, and ventilatory equivalents for O2 and CO2 for both forms of exercise were not significantly different. To determine the optimum exercise test for both treadmill and cycle, we exercised five of the subjects at various work rate increments on both ergometers in a randomized design. The treadmill increments were 0.8, 1.7, 2.5, and 4.2%/min at a constant speed of 3.4 mph, and 1.7 and 4.2%/min at 4.5 mph. Cycle increments were 15, 30, and 60 W/min. The VO2 max was significantly higher on tests where the increment magnitude was large enough to induce test durations of 8-17 min, but the AT was independent of test duration. Thus, for evaluating cardiopulmonary function with incremental exercise testing by either treadmill or cycle, we suggest selecting a work rate increment to bring the subject to the limit of his tolerance in about 10 min.     

The power-duration product--evaluation of a new reference system for cardiopulmonary exercise testing.

The aim of this study was to assess the discriminatory power of the new reference system, power-duration product (PDP), for the analysis of haemodynamic and metabolic variables derived from cardiopulmonary exercise tests. The PDP was calculated as the cumulative index of the product of power (W) times the duration (minutes) of each individual exercise step. The study comprised 30 healthy male volunteers, who were classified into three groups with respect to their regular physical activity: 10 untrained medical students (students), 10 sprinters and long-jumpers (athletes) and 10 endurance athletes performing triathlon (triathletes). Twenty metabolic and haemodynamic variables were recorded throughout exhaustion-limited cycling ergometry. The data were analysed with respect to five reference systems (heart rate, relative and absolute oxygen consumption/body surface area, power, and PDP). A total of 14 differences between modified time courses of haemodynamic and metabolic variables in the three groups of volunteers were observed by reference to PDP, 12 by reference to relative oxygen consumption/body surface area, 11 by reference to heart rate, 8 by reference to absolute oxygen consumption/body surface area, and 7 by reference to power. When using PDP as the reference, the time courses of 8 parameters differed significantly between students and triathletes, 5 between students and athletes, and 1 between athletes and triathletes. In addition to its discriminatory superiority for the comparison of different groups characterized by different cardiopulmonary training and endurance, it was found that PDP permitted a better characterization of the individually performed exercise than the consideration of power per se.     

Doublet stimulation protocol to minimize musculoskeletal stress during paralyzed quadriceps muscle testing.

With long-term electrical stimulation training, paralyzed muscle can serve as an effective load delivery agent for the skeletal system. Muscle adaptations to training, however, will almost certainly outstrip bone adaptations, exposing participants in training protocols to an elevated risk for fracture. Assessing the physiological properties of the chronically paralyzed quadriceps may transmit unacceptably high shear forces to the osteoporotic distal femur. We devised a two-pulse doublet strategy to measure quadriceps physiological properties while minimizing the peak muscle force. The purposes of the study were 1) to determine the repeatability of the doublet stimulation protocol, and 2) to compare this protocol among individuals with and without spinal cord injury (SCI). Eight individuals with SCI and four individuals without SCI underwent testing. The doublet force-frequency relationship shifted to the left after SCI, likely reflecting enhancements in the twitch-to-tetanus ratio known to exist in paralyzed muscle. Posttetanic potentiation occurred to a greater degree in subjects with SCI (20%) than in non-SCI subjects (7%). Potentiation of contractile rate occurred in both subject groups (14% and 23% for SCI and non-SCI, respectively). Normalized contractile speed (rate of force rise, rate of force fall) reflected well-known adaptations of paralyzed muscle toward a fast fatigable muscle. The doublet stimulation strategy provided repeatable and sensitive measurements of muscle force and speed properties that revealed meaningful differences between subjects with and without SCI. Doublet stimulation may offer a unique way to test muscle physiological parameters of the quadriceps in subjects with uncertain musculoskeletal integrity.     

Analysis of heart rate variability during exercise stress testing using respiratory information

This paper presents a novel method for the analysis of heart rate variability (HRV) during exercise stress testing enhanced with respiratory information. The instantaneous frequency and power of the low frequency (LF) and high frequency (HF) bands of the HRV are estimated by parametric decomposition of the instantaneous autocorrelation function (ACF) as a sum of damped sinusoids. The instantaneous ACF is first windowed and filtered to reduce the cross terms. The inclusion of respiratory information is proposed at different stages of the analysis, namely, the design of the filter applied to the instantaneous ACF, the parametric decomposition, and the definition of a dynamic HF band. The performance of the method is evaluated on simulated data as well as on a stress testing database. The simulation results show that the inclusion of respiratory information reduces the estimation error of the amplitude of the HF component from 3.5% to 2.4% in mean and related SD from 3.0% to 1.7% when a tuned time smoothing window is used at an SNR of 15 dB. Results from the stress testing database show that information on respiratory frequency produces HF power estimates which closely resemble those from the simulations which exhibited lower SD. The mean SD of these estimates with respect to their mean trends is reduced by 84% (from 0.74×10⁻³ s⁻² to 0.12×10⁻³ s⁻²). The analysis of HRV in the stress testing database reveals a significant decrease in the power of both the LF and HF components around peak stress.     

The new clinical standard of integrated quadruple stress echocardiography with ABCD protocol

Background

The detection of regional wall motion abnormalities is the cornerstone of stress echocardiography. Today, stress echo shows increasing trends of utilization due to growing concerns for radiation risk, higher cost and stronger environmental impact of competing techniques. However, it has also limitations: underused ability to identify factors of clinical vulnerability outside coronary artery stenosis; operator-dependence; low positivity rate in contemporary populations; intermediate risk associated with a negative test; limited value of wall motion beyond coronary artery disease. Nevertheless, stress echo has potential to adapt to a changing environment and overcome its current limitations.

Integrated-quadruple stress-echo

Four parameters now converge conceptually, logistically, and methodologically in the Integrated Quadruple (IQ)-stress echo. They are: 1- regional wall motion abnormalities; 2-B-lines measured by lung ultrasound; 3-left ventricular contractile reserve assessed as the stress/rest ratio of force (systolic arterial pressure by cuff sphygmomanometer/end-systolic volume from 2D); 4- coronary flow velocity reserve on left anterior descending coronary artery (with color-Doppler guided pulsed wave Doppler). IQ-Stress echo allows a synoptic functional assessment of epicardial coronary artery stenosis (wall motion), lung water (B-lines), myocardial function (left ventricular contractile reserve) and coronary small vessels (coronary flow velocity reserve in mid or distal left anterior descending artery). In “ABCD” protocol, A stands for Asynergy (ischemic vs non-ischemic heart); B for B-lines (wet vs dry lung); C for Contractile reserve (weak vs strong heart); D for Doppler flowmetry (warm vs cold heart, since the hyperemic blood flow increases the local temperature of the myocardium). From the technical (acquisition/analysis) viewpoint and required training, B-lines are the kindergarten, left ventricular contractile reserve the primary (for acquisition) and secondary (for analysis) school, wall motion the university, and coronary flow velocity reserve the PhD program of stress echo.

Conclusion

Stress echo is changing. As an old landline telephone with only one function, yesterday stress echo used one sign (regional wall motion abnormalities) for one patient with coronary artery disease. As a versatile smart-phone with multiple applications, stress echo today uses many signs for different pathophysiological and clinical targets. Large scale effectiveness studies are now in progress in the Stress Echo2020 project with the omnivorous “ABCD” protocol.

     

Development of a new method for testing strength of cells against fluid shear stress

A new method for testing the strength of cells against fluid shear stress by using a long capillary column was proposed. The trajectories of cells in the column were simulated by introducing the Brownian motion model. The Brownian motion was performed by the generation of random numbers. The mean exposure time to shear stress and the mean shear stress acting on the surface of cells were discussed by the result of computer simulation. The mean shear stress acting on the surface of cells flowing in the capillary column was estimated as 4/3-fold of the shear stress at the column wall provided that the ratio of the cell radius to the column radius does not exceed 0.08. The effectiveness of this new method for testing the strength of cells against fluid shear stress was shown.List of Symbols a m radius of cell - c constant - E distribution function - L m length of capillary column - M number of division - N number of division - p probability - Q m³/s flow rate - R m radius of capillary column - r m radial position - t s time - T s exposure time - T_m s mean exposure time - T₀ s mean residence time - m/s axial velocity - u_mm/s cross-sectional flow velocity - z m axial position - s^–1 shear rate - _ws^–1 shear rate at wall - Pa s viscosity - spherical coordinate - spherical coordinate - Pa shear stress - _mPa mean shear stress - _wPa shear stress at wall     

Reliability of an exercise protocol designed to evaluate resistance exercise performance

The purpose of the present investigation was to evaluate the reliability a protocol used to assess short-term resistance exercise performance. Six men participated in this investigation after giving their consent. Subjects (N = 6) performed 6 sets of leg extensions at 80% of 10 repetition maximum (RM). Ten repetitions were performed during the first 3 sets; during the last 3 sets subjects exercised to fatigue. Ninety seconds of seated passive recovery separated each set. Subsequently, 2 experimental trials were conducted in which the exercise protocol was identical to the familiarization trial. There was a significant decline in performance from set 4 (13.5 +/- 0.9 reps) to set 5 (11.9 +/- 0.8 reps) and set 4 to set 6 (10.8 +/- 1.0 reps), suggesting that the protocol did induce fatigue. The intraclass correlations were 0.992, 0.992, and 0.993 for the fourth, fifth, and sixth sets, respectively. The average coefficients of variation for the fourth, fifth, and sixth sets were 6.7, 2.7, and 7.1%, respectively. These data suggest that the resistance training protocol used in this investigation is reliable and may be useful in evaluating interventions designed to improve fatigue resistance.     

A standardized protocol for repeated social defeat stress in mice

A major impediment to novel drug development has been the paucity of animal models that accurately reflect symptoms of affective disorders. In animal models, prolonged social stress has proven to be useful in understanding the molecular mechanisms underlying affective-like disorders. When considering experimental approaches for studying depression, social defeat stress, in particular, has been shown to have excellent etiological, predictive, discriminative and face validity. Described here is a protocol whereby C57BL/6J mice that are repeatedly subjected to bouts of social defeat by a larger and aggressive CD-1 mouse results in the development of a clear depressive-like syndrome, characterized by enduring deficits in social interactions. Specifically, the protocol consists of three important stages, beginning with the selection of aggressive CD-1 mice, followed by agonistic social confrontations between the CD-1 and C57BL/6J mice, and concluding with the confirmation of social avoidance in subordinate C57BL/6J mice. The automated detection of social avoidance allows a marked increase in throughput, reproducibility and quantitative analysis. This protocol is highly adaptable, but in its most common form it requires 3-4 weeks for completion.     

A new method for testing gaussian processes

A method of testing gaussian processes, based on the characteristic function, is presented. An evaluation of possible gaussian inclusions in some stochastic processes is also proposed.     

Evaluation of a microprocessor-controlled exercise testing system

We evaluated a new exercise-testing system (Beckman Horizon MMC), incorporating a microprocessor that controls the acquisition of data, corrects for time delays, applies calibration factors, ensures quality control, and presents results in a variety of formats. Precision of measurements of ventilation (VE) and mixed expired gas concentrations was high. In steady-state exercise (n = 100) VO2 was measured with a precision (+/- SD) of 66 ml/min (4.3%), (r = 0.991); there was a small (4.62%) systematic underestimation of VCO2, but precision was comparable with VO2, with SD being 67 ml/min (4.55%) (r = 0.993). Good agreement was obtained between measurements made in progressive incremental exercise in healthy subjects with correlation coefficients of 0.997 for VE, 0.995 for VO2, and 0.994 for VCO2. Agreement in patients with cardiorespiratory disorders (n = 10) was similar, except in three patients in whom a variable pattern of breathing limited strict comparisons. Comparison with a breath-by-breath analysis system (n = 5) showed that rapid changes in VE, VCO2, and VO2 were followed accurately; the half time for a change in VO2 was not systematically different between the two systems (SD, 3.34 s, r = 0.951). The incorporation of microprocessor-controlled calibration procedures, which are simple to carry out frequently, was judged to be an important feature of this system.     

DINC: A new AutoDock-based protocol for docking large ligands

Background

Using the popular program AutoDock, computer-aided docking of small ligands with 6 or fewer rotatable bonds, is reasonably fast and accurate. However, docking large ligands using AutoDock's recommended standard docking protocol is less accurate and computationally slow.

Results

In our earlier work, we presented a novel AutoDock-based incremental protocol (DINC) that addresses the limitations of AutoDock's standard protocol by enabling improved docking of large ligands. Instead of docking a large ligand to a target protein in one single step as done in the standard protocol, our protocol docks the large ligand in increments. In this paper, we present three detailed examples of docking using DINC and compare the docking results with those obtained using AutoDock's standard protocol. We summarize the docking results from an extended docking study that was done on 73 protein-ligand complexes comprised of large ligands. We demonstrate not only that DINC is up to 2 orders of magnitude faster than AutoDock's standard protocol, but that it also achieves the speed-up without sacrificing docking accuracy. We also show that positional restraints can be applied to the large ligand using DINC: this is useful when computing a docked conformation of the ligand. Finally, we introduce a webserver for docking large ligands using DINC.

Conclusions

Docking large ligands using DINC is significantly faster than AutoDock's standard protocol without any loss of accuracy. Therefore, DINC could be used as an alternative protocol for docking large ligands. DINC has been implemented as a webserver and is available at http://dinc.kavrakilab.org. Applications such as therapeutic drug design, rational vaccine design, and others involving large ligands could benefit from DINC and its webserver implementation.