In vivo measurement of protein functional changes

Conformational changes in proteins are fundamental to all biological functions. In protein science, the concept of protein flexibility is widely used to describe protein dynamics and thermodynamic properties that control protein conformational changes. In this study, we show that urea, which has strong sedative potency, can be administered to fish at high concentrations, and that protein functional changes related to anesthesia induction can be measured in vivo. Ctenopharyngodon idellus (the grass carp) has two different types of N-methyl d-aspartate (NMDA) receptors, urea-insensitive and urea-sensitive, which are responsible for the heat endurance of fish. The urea-sensitive NMDA receptor showed high protein flexibility, the gamma aminobutyric acid (GABA) receptor showed less flexibility, and the protein that is responsible for ethanol anesthesia showed the lowest flexibility. The results suggest that an increase in protein flexibility underlies the fundamental biophysical mechanisms of volatile general anesthetics.     

Effect of the intra-abdominal pressure and the center of segmental body mass on the lumbar spine mechanics - a computational parametric study

Determination of physiological loads in human lumbar spine is critical for understanding the mechanisms of lumbar diseases and for designing surgical treatments. Computational models have been used widely to estimate the physiological loads of the spine during simulated functional activities. However, various assumptions on physiological factors such as the intra-abdominal pressure (IAP), centers of mass (COMs) of the upper body and lumbar segments, and vertebral centers of rotation (CORs) have been made in modeling techniques. Systematic knowledge of how these assumptions will affect the predicted spinal biomechanics is important for improving the simulation accuracy. In this paper, we developed a 3D subject-specific numerical model of the lumbosacral spine including T12 and 90 muscles. The effects of the IAP magnitude and COMs locations on the COR of each motion segment and on the joint/muscle forces were investigated using a global convergence optimization procedure when the subject was in a weight bearing standing position. The data indicated that the line connecting the CORs showed a smaller curvature than the lordosis of the lumbar spine in standing posture when the IAP was 0?kPa and the COMs were 10?mm anterior to the geometric center of the T12 vertebra. Increasing the IAP from 0 kPa to 10 kPa shifted the location of CORs toward the posterior direction (from 1.4?±?8.9 mm anterior to intervertebral disc (IVD) centers to 40.5?±?3.1 mm posterior to the IVD centers) and reduced the average joint force (from 0.78?±?0.11 Body weight (BW) to 0.31?±?0.07 BW) and overall muscle force (from 349.3?±?57.7 N to 221.5?±?84.2 N). Anterior movement of the COMs from -30 mm to 70 mm relative to the geometric center of T12 vertebra caused an anterior shift of the CORs (from 25.1?±?8.3 mm posterior to IVD centers to 7.8?±?6.2 mm anterior to IVD centers) and increases of average joint forces (from 0.78?±?0.1 BW to 0.93?±?0.1 BW) and muscle force (from 348.9?±?47.7 N to 452.9?±?58.6 N). Therefore, it is important to consider the IAP and correct COMs in order to accurately simulate human spine biomechanics. The method and results of this study could be useful for designing prevention strategies of spinal injuries and recurrences, and for enhancing rehabilitation efficiency.     

In vivo measurement of phytochrome in tomato fruit

Presence of phytochrome in two kinds of tomatoes (Lycopersicon esculentum Mill.), the yellow lutescent strain and cherry tomatoes (L. esculentum Mill. var. cerasiformecv. Red Cherry), was established by measuring the absorption difference spectra of the whole fruit after irradiation with red and with far red light. Phytochrome content was determined in yellow lutescent tomatoes and decreased gradually during the ripening period.     

In vivo measurement of translational stiffness of rabbit knees

This paper describes the design, evaluation, and preliminary results of a specialized testing device and surgical protocol to determine translational stiffness of a rabbit knee, replicating the clinical anterior drawer test. Coronal-plane transverse pins are inserted through the rabbit leg, two in the tibia and one in the distal femur, to hold and reproducibly position the leg in the device for tests at multiple time points. A linear stepper motor draws the tibia upward then returns to the home position, and a load cell measures the resisting force; force-displacement knee stiffness is then calculated. Initial evaluation of this testing device determined the effects of preconditioning, intra-operator repeatability, rabbit-to-rabbit variability, knee flexion angle (90 degrees vs. 135 degrees ), and anterior cruciate ligament (ACL) sectioning (0%, 25%, 50%, 75%, 100%). Knee stiffness generally decreased as ACL sectioning increased. This testing device and surgical protocol provide an objective and efficient method of determining translational rabbit knee stiffness in vivo, and are being used in an ongoing study to evaluate the effect of knee instability (via partial to complete ACL sectioning) on the development of post-traumatic osteoarthritis.     

In vivo tracking of the human patella.

The purpose of this study was to describe the dynamic, in vivo, three-dimensional tracking pattern of the patella for one normal male subject. Intracortical pins were inserted into the patella, tibia, and femur. The subject performed seated and squatting knee flexion/extension, and maximum voluntary quadriceps contractions. In addition, the vastus medialis oblique was subjected to maximal electrical stimulation. Motions of the markers attached to the intracortical pins were analyzed using an automated video system. Patellar and tibial motions were determined relative to a femoral reference system. While the tibia flexed 50 degrees from full extension (seated condition), the patella flexed 30.3 degrees, tilted laterally 10.3 degrees, and shifted laterally 8.6 mm. In general, these results show qualitative agreement with the data collected from cadaveric specimens [van Kampen and Huiskes, J. orthop. Res. 8, 372-382 (1990)]. The differences present may reflect different passive constraints to patellar motions, and different relative loading of the individual quadriceps components, in our study compared to the cadaveric study. Only small differences were found between patellar motions in the seated and squatting conditions. Differences in patellar displacements produced by (1) maximal electrical stimulation of the vastus medialis oblique, and (2) maximum voluntary quadriceps contraction, at 30 degrees knee flexion and full extension, may reflect the dominant influence of passive constraints, and the vastus lateralis, on normal patellar motions. Further in vivo study of patellar tracking seems warranted to evaluate surgical and conservative interventions for patellofemoral disorders.     

In vivo effect of copper status on cisplatin-induced nephrotoxicity

Cisplatin is a widely used antitumor agent; however, tumor resistance and severe side effects limit its use. It is well accepted that cisplatin toxicity can be modulated in vitro in cell cultures by copper salts. In the present work, mice with different blood serum copper status were treated with a single intraperitoneal cisplatin injection at a dose of 5 mg/kg, monitored for 3 days in metabolic cages and analyzed for renal function. Both copper-deficient and copper-overloaded mice displayed more severe early proteinuria and retarded platinum excretion than control mice. The effects of copper status on cisplatin-induced nephrotoxicity are discussed.     

In vivo measurement of bone aluminium: recent developments

A biomarker of aluminium accumulation in the human body can play a valuable role in determining health effects of chronic aluminium exposure, complementing other human and environmental monitoring data. In vivo neutron activation provides such a non-invasive biomarker. To date, the best in vivo neutron activation system used thermalised neutrons from a nuclear reactor at Brookhaven National Laboratory, which suffered only slightly from interference from other elements, primarily phosphorus, and from the disadvantage of restricted accessibility. At McMaster, we use a nuclear reaction on an accelerator to select neutron energy, which eliminates the interferences. Spectral decomposition analysis improved sensitivity. A new 4pi detection system also enhanced sensitivity. Together these improvements yield a minimum detection limit of 0.24 mgAl in a hand, slightly better than at Brookhaven and equivalent to "normal" levels. Further improvements should result from a new irradiation cavity and from using a higher proton current on the accelerator to shorten irradiation times. The system is now ready for pilot human studies.     

In vivo measurement of bending stiffness in fracture healing

Background  

Measurement of the bending stiffness a healing fracture represents a valid variable in the assessment of fracture healing. However, currently available methods typically have high measurement errors, even for mild pin loosening. Furthermore, these methods cannot provide actual values of bending stiffness, which precludes comparisons among individual fractures. Thus, even today, little information is available with regards to the fracture healing pattern with respect to actual values of bending stiffness. Our goals were, therefore: to develop a measurement device that would allow accurate and sensitive measurement of bending stiffness, even in the presence of mild pin loosening; to describe the course of healing in individual fractures; and help to evaluate whether the individual pattern of bending stiffness can be predicted at an early stage of healing.     

In vivo measurement of muscle output in intact Drosophila

We describe our methods for analysing muscle function in a whole intact small insect, taking advantage of a simple flexible optical beam to produce an inexpensive transducer with wide application. We review our previous data measuring the response to a single action potential driven muscle twitch to explore jumping behaviour in Drosophila melanogaster. In the fruitfly, where the sophisticated and powerful genetic toolbox is being widely employed to investigate neuromuscular function, we further demonstrate the use of the apparatus to analyse in detail, within whole flies, neuronal and muscle mutations affecting activation of muscle contraction in the jump muscle. We have now extended the use of the apparatus to record the muscle forces during larval and other aspects of adult locomotion. The robustness, simplicity and versatility of the apparatus are key to these measurements.     

In vivo measurement of glucose concentration with lasers.

Common polarimeters defined the concentration of an optically active sample by measuring the intensity of the light beam after it has passed through the analyser. Consequently, it can be stated that up to now, there have been the disadvantages of having a signal dependent on absolute light intensity. The new method that is described has a sensitivity that is in great measure independent of absolute light intensity, whereby only one light trace is necessary. The new principle uses no mechanical rotations. Instead, an electrical signal indicates the amount of optical rotation of the sample. The high sensitivity that can be reached is theoretically only limited by polarization noise. By going to the uttermost physical and electronic lengths, sensitivity values of more than 10(-5) degrees can be reached. Furthermore, the mechanical dimensions of the apparatus can be made very small by the application of a solid-state laser.     

Method for in vivo measurement of intraosseous pressure of the patella]

During the development of degenerative joint disease (osteoarthritis, chondropathy), a diagnostic or even pathogenetic role is attributed to the phenomenon of intraosseous pressure (IOP). Owing to technical problems and a lack of systematic experimental or clinical studies on the control mechanisms of the IOP, the actual importance of this factor is still not known with certainty. Now, a measuring method that enables correct recording of the IOP and standardized on-line processing of the measured signal minimize artefact-related problems. The technique is evaluated for reliability in an in vitro model of the human patella and in a limited clinical study of the IOP in patients undergoing knee surgery for various reasons. Factors such as intra-articular effusion, joint position or changes in intra-articular soft tissue are examined for their influence of the primary signal.     

In vivo 3-dimensional measurement of the force exerted on a tooth during clenching

We have developed a three-dimensional (3D) force-measuring device for teeth and used it to measure functional forces in vivo. It comprises an inner part forming a metal core (abutment), a 3D piezoelectric force transducer, and an outer part forming a metal crown, all joined together with a steel screw. The force transducer can measure +/- 500 N along the z-axis and +/- 150 N along the x- and y-axes. We evaluated the relationship between output and load and the effects of hysteresis and temperature on the output. The transducer had high linearity (r>0.9999), low hysteresis (1.7% at maximum), and high thermal stability (0.05% per degree) along each axis. The measuring device was mounted on the maxillary left second molar of a healthy male subject; the tooth had been endodontically treated (neurovascular bundle removed) and prepared for metal abutment and a crown. The 3D load calculated from the outputs of the transducer was expressed as a vector of the coordinates based on the Frankfort horizontal (x-y) and sagittal (y-z) planes. The force measured during maximum voluntary clenching was about 170 N; the force vector was directed from the crown to the root medially at an angle of about 10 degrees from the y-z plane and posteriorly at an angle of about 3 degrees from the x-z plane. This transducer will enable measurement of forces applied to different types of prosthetic appliances and has the potential to provide important basic in vivo data for analysis using computer simulation.     

In vivo O2 measurement inside single photosynthetic cells

The oxygen evolution of single cells was investigated using a nano-probe with an ultra-micro electrode (UME) in a submicron sized system in combination with a micro-fluidic system. A single cell was immobilized in the micro-fluidic system and a nano-probe was inserted into the cytosolic space of the single cell. Then, the UME was used for an in vivo amperometric experiment at a fixed potential and electrochemical impedance spectroscopy to detect oxygen evolution of the single cell under various light intensities.     

In vivo simulation of human pharmacokinetics in the rabbit

The evaluation of drugs in vivo is often based on experimental models using small animals such as mice, rats and rabbits. However, these models could be improved to correspond more closely to the human situation if the pharmacokinetics of the drugs tested in animals were similar to that observed in humans. The use of a computer-controlled pump allowing an adequate flow of tobramycin and amikacin to be infused into rabbits enabled us to simulate the human pharmacokinetics of these antibiotics in vivo in this study. The function defining the rate of infusion required to perform the simulation of an intravenous bolus was first determined generally and symbolically for linear pharmacokinetic models independently from the number of compartments involved. The practical simulation of a decreasing monoexponential serum profile with a half-life of 2 h (one-compartment model for the human pharmacokinetics of aminoglycosides) was then studied for tobramycin and amikacin on the basis of a two-compartment model in the animal. The kinetics obtained had an apparent elimination half-life of 1.97 and 1.86 h, respectively. Linearity of the semilogarithmic regressions of the profiles obtained was quite sound. Finally, an a posteriori analysis of the pharmacokinetic model and its parameters is proposed on the basis of the results obtained after simulation.