Jeung Hwan Choi

Cellular Biophysics During Freezing of Rat and Mouse Sperm Predicts Post-thaw Motility

Though cryopreservation of mouse sperm yields good survival and motility after thawing, cryopreservation of rat sperm remains a challenge. This study was designed to evaluate the biophysics (membrane permeability) of rat in comparison to mouse to better understand the cooling rate response that contributes to cryopreservation success or failure in these two sperm types. In order to extract subzero membrane hydraulic permeability in the presence of ice, a differential scanning calorimeter (DSC) method was used. By analyzing rat and mouse sperm frozen at 5°C/min and 20°C/min, heat release signatures characteristic of each sperm type were obtained and correlated to cellular dehydration. The dehydration response was then fit to a model of cellular water transport (dehydration) by adjusting cell-specific biophysical (membrane hydraulic permeability) parameters Lpg and ELp. A “combined fit” (to 5°C/min and 20°C/min data) for rat sperm in Biggers-Whitten-Whittingham media yielded Lpg = 0.007 μm min−1 atm−1 and ELp = 17.8 kcal/mol, and in egg yolk cryopreservation media yielded Lpg = 0.005 μm min−1 atm−1 and ELp = 14.3 kcal/mol. These parameters, especially the activation energy, were found to be lower than previously published parameters for mouse sperm. In addition, the biophysical responses in mouse and rat sperm were shown to depend on the constituents of the cryopreservation media, in particular egg yolk and glycerol. Using these parameters, optimal cooling rates for cryopreservation were predicted for each sperm based on a criteria of 5%–15% normalized cell water at −30°C during freezing in cryopreservation media. These predicted rates range from 53°C/min to 70°C/min and from 28°C/min to 36°C/min in rat and mouse, respectively. These predictions were validated by comparison to experimentally determined cryopreservation outcomes, in this case based on motility. Maximum motility was obtained with freezing rates between 50°C/min and 80°C/min for rat and at 20°C/min with a sharp drop at 50°C/min for mouse. In summary, DSC experiments on mouse and rat sperm yielded a difference in membrane permeability parameters in the two sperm types that, when implemented in a biophysical model of water transport, reasonably predict different optimal cooling rate outcomes for each sperm after cryopreservation.

A quantitative analysis of the thermal properties of porcine liver with glycerol at subzero and cryogenic temperatures

There is a lack of information on the effect of cryoprotective agents (CPAs) on the thermal properties of biomaterials at cryobiologically relevant temperatures (i.e. <233.15K, -40 degrees C). Thermal properties that are of most interest include: thermal conductivity, density, specific heat, and latent heat resulting from phase change in tissue systems. Availability of such information would be beneficial for accurate mathematical modeling of cryobiological applications. Recently, we reported these thermal properties in phosphate buffered saline (PBS) with varying concentrations of glycerol, a widely used cryoprotective agent. In this study we extend these results by assessing the effects of glycerol on the thermal properties of porcine liver at subzero temperatures. Differential scanning calorimeter (DSC) was used to measure the specific heat and the latent heat release of porcine liver immersed in PBS and varying concentrations of glycerol. The specific heat data obtained from the DSC experiments were also used to predict the bulk thermal conductivity. This was done using a transient heat transfer model with a thermistor probe technique. Results show that the introduction of glycerol significantly alters thermal properties from known values for H2O and non-treated liver. Therefore, inaccuracies in thermal predictions can be expected due to the application of measured vs. predicted thermal properties such as from weight averaging. This supports the need for these and other measurements of biomaterial thermal properties, with and without CPA addition, in the cryogenic regime.

A Hydrophobic Gel Phantom for Study of Thermochemical Ablation: Initial Results Using a Weak Acid and Weak Base

PURPOSE To develop a model for study of exothermic chemical reactions potentially useful for tissue ablation. MATERIALS AND METHODS Seven gelatins ranging from 0.5% to 30% wt/vol with and without 15% or 30% caps and several commercial gels were evaluated. Baseline temperature measurements were taken. Acetic acid and ammonium hydroxide were sequentially injected over periods of 10-15 seconds in 1-mL aliquots, forming a discrete aqueous reaction chamber. Congo red pH indicator was included to assess the reaction. A thermocouple allowed data collection at completion of injection and every 15 seconds for 5 minutes. Injections were performed in triplicate, and average temperatures for each time point were reported. RESULTS Gelatins fractured or refluxed even at the lowest concentrations tested. Most commercial gels proved too viscous and likewise led to reflux along the needle tract. A mineral oil-based gel was selected because of its ability to form a chamber without reflux or fracture and its clear colorless character, hydrophobic nature, chemical stability, viscosity, specific gravity, and cost. Temperatures during the first 60 seconds of the neutralization reaction showed an immediate increase that correlated well with concentration. CONCLUSIONS The oil gel phantom is a safe, useful, readily available, inexpensive model to study mixing behaviors and maximum heating potentials for reactions that may prove useful in thermochemical tissue ablation for oncologic interventions. Measurable temperature changes occurred even at the lowest concentrations, and higher concentrations produced a greater release of heat energy.

A simple transient method for measurement of thermal conductivity of rigid polyurethane foams

Rigid polyurethane foams (PU) are widely used as thermal insulators in various applications. The thermal conductivity of the foam is the key parameter that governs the efficiency of thermal insulation provided by the foam. The usual technique employed to measure thermal conductivity is based on the rate of steady state heat transfer across a known thickness, induced by two different known temperatures at two opposite surfaces of the foam. We introduce a technique based on the transient measurement of heat transfer measured by an embedded needle probe. This technique is not only rapid but the instrumentation required for such a measurement is simple and the cost is only a fraction of the steady state counterpart. The values of thermal conductivity obtained by both methods are compared and found to agree within 4% over the range of 0.02—0.03 W/mK, which is the usual range of thermal conductivity for commercial rigid PU foams. The sensitivity of the needle probe technique is demonstrated by measuring the thermal conductivity values of foams made with various concentrations of chemical blowing agent (water). The present technique is also shown to be effective for measuring the thermal conductivity of small samples, especially, free rise cup foams for which the steady state technique can not be used.

Mapping tissue oxygen in vivo by photoacoustic lifetime imaging

Oxygen plays a key role in the energy metabolism of living organisms. Any imbalance in the oxygen levels will affect the metabolic homeostasis and lead to pathophysiological diseases. Hypoxia, a status of low tissue oxygen, is a key factor in tumor biology as it is highly prominent in tumor tissues. However, clinical tools for assessing tissue oxygenation are limited. The gold standard is polarographic needle electrode which is invasive and not capable of mapping (imaging) the oxygen content in tissue. We applied the method of photoacoustic lifetime imaging (PALI) of oxygen-sensitive dye to small animal tissue hypoxia research. PALI is new technology for direct, non-invasive imaging of oxygen. The technique is based on mapping the oxygen-dependent transient optical absorption of Methylene Blue (MB) by pump-probe photoacoustic imaging. Our studies show the feasibility of imaging of dissolved oxygen distribution in phantoms. In vivo experiments demonstrate that the hypoxia region is consistent with the site of subcutaneously xenografted prostate tumor in mice with adequate spatial resolution and penetration depth.

Effects of glycerol on the thermal properties of phosphate buffered saline and porcine liver at subzero temperatures

Improvements in the prediction of thermal behavior during cryosurgery and cryopreservation can help improve the outcome of these cryobiological applications. The accuracy of the models depends on numerous factors including the kinetics and energy release during phase change phenomena and knowledge of thermal properties. Furthermore, connecting the thermal properties to crystalline, amorphous, and other phases adds an important mechanistic dimension that can also improve and direct an outcome. However, insufficient data for thermal properties in the subzero domain result in reliance on property estimations based usually upon tabulated water-ice data or weight averaged values from known materials primarily in temperature ranges above −40 °C [1]. This study focused on expanding the thermal properties database for both solutions and tissues. Results for Phosphate Buffered Saline (PBS) and porcine liver with glycerol at subzero temperatures (−150 ∼ 0 °C) are reported. The shifting of thermal property values due to sample crystallization, amorphous phase transition, and melting is discussed.© 2007 ASME