A pig model of hepatic cryotherapy. In vivo temperature distribution during freezing and histopathological changes

We aimed to assess the temperature distribution in the cryolesion during hepatic cryotherapy and the association with postoperative histological changes to optimise the technique and allow better preoperative planning. Hepatic cryolesions were produced in 22 pigs following laparotomy using a CMS-cryosystem and 8mm-AccuProbe-Cryoprobes. The temperature was measured in 1 min intervals at different distances from the probe during freezing. The animals were treated in 5 groups: (i) single freezing of 20 min; (ii) double freezing of 20 min each; (iii) single freezing of 40 min; (iv) single freezing of 20 min (n=4), histology at 1 week p.o., and (v) single freezing of 20 min and Pringle manoeuvre; [(i)-(iii) and (v): histology at 24 h p.o.]. The mean diameter of the -38 degrees C isotherm, i.e., the zone of effective treatment for colorectal metastases was 37 mm for group (i) with a mean iceball diameter of 59 mm and about 46 mm for groups (ii, iii, and v) with mean iceball diameters of 78, 75, and 75 mm, respectively. At 7 days postoperatively secondary necrosis was seen in the largest central part of the lesion, wherever temperatures of -15 degrees C or lower were achieved during cryosurgery. Under the hypothesis that -38 degrees C is the effective temperature for the destruction of colorectal liver metastases, a lesion of 37-mm diameter may be effectively treated with a single 8mm-AccuProbe-Cryoprobe and a 20 min single freeze cycle and a lesion of 46 mm may be effectively treated when a double freeze-thaw cycle of 20 min each, a single freeze cycle of 40 min, or a 20 min single freeze cycle with additional Pringle manoeuvre is used, when it is perfectly placed in the lesion.     

Advanced hepatic tissue destruction in ablative cryosurgery: potentials of intermittent freezing and selective vascular inflow occlusion

Recent studies indicate that cryosurgery represents a promising approach to treat non-resectable liver tumors. To improve parenchymal tissue destruction, a variety of modifications of the freeze-thaw procedure have been suggested, including repetitive freezing and portal-triad cross-clamping. The aim of the present study was to analyze whether intermittent freezing by application of a double freeze-thaw procedure or selective vascular inflow occlusion are more effective than a single freeze-thaw cycle to achieve complete hepatic tissue destruction. Using a porcine model, intrahepatic cryolesions were induced by freezing the hepatic tissue for a total of 15 min (n=6, SF). Additional animals (n=6) underwent a double freeze-thaw cycle of 7.5 min each (DF). A third group of animals (n=6) was treated by a single 15-min freeze-thaw cycle during selective vascular inflow occlusion (VO-SF). Seven days after freezing, DF did not change the volume of the cryolesion (25.4+/-1.7 cm(3)) compared to SF (29.9+/-3.7 cm(3)), however, resulted in enhanced destruction of hepatocyte nuclear morphology (DF-score: 2.4+/-0.2 versus SF-score: 1.1+/-0.3; p<0.05) and attenuated leukocyte infiltration within the margin of the cryolesion (DF-score: 1.5+/-0.2 versus SF-score: 2.8+/-0.1; p<0.05). VO-SF was also effective to significantly enhance destruction of hepatocyte nuclear morphology (2.8+/-0.1; p<0.05 versus SF), but, additionally, markedly increased the volume of the cryolesions (43.3+/-5.3 cm(3); p<0.05 versus SF and DF). Interestingly, VO-SF further increased the number of apoptotic cells, while leukocyte infiltration (2.3+/-0.3) was not affected compared to that after SF-treatment. Thus, our data indicate that both DF and VO-SF are effective to enhance parenchymal cell destruction within the margin of the cryolesion. VO-SF additionally increases the volume of the lesion and may therefore be most attractive for successful clinical application.     

Cryosurgical changes in the porcine kidney: histologic analysis with thermal history correlation

Advances in minimally invasive renal cryosurgery have renewed interest in the relative contributions of direct cryothermic and secondary vascular injury-associated ischemic cell injury. Prior studies have evaluated renal cryolesions seven or more days post-ablation and postulated that vascular injury is the primary cell injury mechanism; however, the contributions of direct versus secondary cell injury are not morphologically distinguishable during the healing/repair stage of a cryolesion. While more optimal to evaluate this issue, minimal acute (< or = 3 days) post-ablation histologic data with thermal history correlation exists. This study evaluates three groups of porcine renal cryolesions: Group (1) in vitro non-perfused (n = 5); Group (2) in vivo 2-h post-ablation perfused (n = 5); and Group (3) in vivo 3-day post-ablation perfused (n = 6). The 3.4 mm argon-cooled cryoprobe's thermal history included a 75 degrees C/min cooling rate, -130 degrees C end temperature, 60 degrees C/min thawing rate, and 15-min freeze time. An enthalpy-based mathematical model with a 2-D transient axisymmetric numerical solution with blood flow consideration was used to determine the thermal history within the ice ball. All three groups of cryolesions showed histologically similar central regions of complete cell death (CD) and transition zones of incomplete cell death (TZ). The CD had radii of 1.4, 1.1, and 1.0 cm in the non-perfused, 2-h and 3-day lesions, respectively. Capillary thrombosis was present in the 2-h perfused cryolesions with the addition of TZ arteriolar/venous thrombosis in the 3-day perfused lesions. Thermal modeling revealed the outer CD boundary in all three groups experienced similar thermal histories with an approximately -20 degrees C end temperature and 2 degrees C/min cooling and thawing rates. The presence of similar CD histology and in vitro/in vivo thermal histories in each group suggests that direct cryothermic cell injury, prior to or synchronous with vascular thrombosis, is a primary mediator of cell death in renal cryolesions.     

Pre-treatment inflammation induced by TNF-alpha augments cryosurgical injury on human prostate cancer

Vascular injury is a major mechanism of cryosurgical destruction. The extent of vascular injury may be affected by the addition of molecular adjuvants. This study, in addition to determining the injury mechanism in the LNCaP Pro 5 human prostate cancer subline grown in a nude mouse, examined the effect of cytokine TNF-alpha on cryosurgery of an in vivo microvascular preparation (Dorsal Skin Flap Chamber). A comparison of injury data to a thermal model indicated that the minimum temperature after moderate cooling, thawing, and hold time required for causing necrosis was 3.5+/-6.9 degrees C in TNF-alpha-treated LNCaP Pro 5 tumor tissue (n=4) and -9.8+/-5.8 degrees C in TNF-alpha-treated normal skin of the nude mouse (n=4). Compared to tissues without TNF-alpha treatment, where the minimum temperature required for causing necrosis was -16.5+/-4.3 degrees C in LNCaP Pro 5 tumor tissue (n=8) and -24.4+/-7.0 degrees C in normal skin of the nude mouse (n=9), the results indicate the local use of TNF-alpha can dramatically increase the threshold temperature of cryo-destruction by more than 10 degrees C (p <0.01). These findings were consistent with the hypothesis that vascular-mediated injury is responsible for defining the edge of the cryolesion in microvascular-perfused tissue, and therefore pre-induced inflammation can augment cryoinjury. The local use of TNF-alpha to pre-inflame prostate cancer promises to increase both the ability of freezing to destroy cancer as well as improve the ability of ultrasound or other iceball-monitoring techniques to predict the outcome of the treatment.     

Parametric study of radiofrequency ablation in the clinical practice with the use of two-compartment numerical models

The objective of the current work was to simulate radiofrequency ablation (RFA) with theoretical and realistic computational models, which correspond to single-compartment models and clinical scenarios. A 3D model in a cubic region of 12 cm edge was studied representing either a homogeneous model or real clinical scenarios in three human tissues, i.e., liver, lung and kidney. An active electrode was placed at the center of the model. Various tumor sizes (1–3 cm) and source voltages (10–30 V) were investigated for the second case of a two-compartment model. In the case of a 3-cm tumor in diameter, the electrical and thermal problems (at steady state) were solved to calculate the temperature distribution within the tumor and tissue. Lesion volume was quantified using the Arrhenius equation and the isothermals of 50 and 60 °C. The physical properties of all materials were constant during the simulations, i.e., no changes with temperature were considered. It was found that tumor conductivity was low to achieve significant damage in the tumor; in all clinical scenarios, saline-enhanced RFA was necessary and led to a more efficient tumor destruction. It was also shown that highly perfused tissues, such as liver and kidney, block the energy deposition within them, in contrast to lung, and, thus, require a further saline enhancement. Finally, the effect of perfusion on lesion size was studied, and it was concluded that tumor perfusion was more significant than surrounding tissue perfusion.     

An assessment of tumor cell viability after in vitro freezing

The identification of the minimum lethal temperature for tumor cells in vivo is difficult because of the secondary factors that are associated with the cryoinjury. This study attempts to identify this temperature by a combination of in vitro and in vivo techniques. Suspensions of Walker carcinoma cells were frozen at a rate of 1 degree C/min without cryoprotection, to either -10, -15, -20, -25, -30, -35 or -40 degrees C and held at that temperature for either 0, 10, 20, or 30 min. After spontaneous rewarming viability was assessed by a combination of vital dye studies and the growth of tumor cells inoculated into the liver and subcutaneous tissue of male, Sprague-Dawley rats. Trypan blue studies indicated that less than 1% of the cells frozen to -35 degrees C were considered viable, yet significant tumor take rates were noted, suggesting that for some cells the cryoinjury is reversible. As expected tumor take rates were reduced by lowering the temperature but were independent of the holding time. The volume doubling time and final tumor volume of the subcutaneous tumors was similar to that of controls, indicating that the growth potential of the cells which survive freezing is normal. The minimum lethal temperature was dependent upon the site of inoculation, subcutaneous tumors developing from cells frozen to -35 degrees C, whereas liver tumors did not develop from cells frozen beyond -25 degrees C, this may have important clinical implications.     

Thermomechanical analysis of collagen crosslinking in the developing lamb pericardium

In a developing lamb model, we have used hydrothermal isometric tension (HIT) techniques to assess collagen crosslink stability and its contribution to the mechanical properties of the pericardium. Strip samples of tissue were either: (i) heated to a 90 degrees C isotherm or (ii) heated in 5 degrees C increments between 60-90 degrees C and then 93 and 97 degrees C isotherms. The half-life of stress relaxation associated with peptide bond hydrolysis (t1/2) was calculated at each isotherm. The activation energy, Eact, for the hydrolysis-associated relaxation process was also calculated using the data from the stepwise HIT tests--a technical improvement which significantly reduces the experimental time required to develop statistically valid measurements. Crosslinking in the pericardium increased during development and was demonstrated both by thermoelasticity and by resistance to enzymatic solubilization. We observed greater conversation to thermally stable crosslinks upon maturation, the ratio of the NaBH4-stabilized/unstabilized half-lives peaking at 21 days postnatal. Whereas tissue from lambs (119 day fetal, and 3 day and 21 day postpartum) showed an early maximum and rapid decay of force, NaBH4 stabilization significantly increased thermal stability and yielded profiles similar to those in adult tissue.     

Cryosurgery of normal and tumor tissue in the dorsal skin flap chamber: Part II--injury response

It has been hypothesized that vascular injury may be an important mechanism of cryosurgical destruction in addition to direct cellular destruction. In this study we report correlation of tissue and vascular injury after cryosurgery to the temperature history during cryosurgery in an in vivo microvascular preparation. The dorsal skin flap chamber implanted in the Copenhagen rat, was chosen as the cryosurgical model. Cryosurgery was performed in the chamber on either normal skin or tumor tissue propagated from an AT-1 Dunning rat prostate tumor, as described in a companion paper (Hoffmann and Bischof, 2001). The vasculature was then viewed at 3 and 7 days after cryoinjury under brightfield and FITC-labeled dextran contrast enhancement to assess the vascular injury. The results showed that there was complete destruction of the vasculature in the center of the lesion and a gradual return to normal patency moving radially outward. Histologic examination showed a band of inflammation near the edge of a large necrotic region at both 3 and 7 days after cryosurgery. The area of vascular injury observed with FITC-labeled dextran quantitatively corresponded to the area of necrosis observed in histologic section, and the size of the lesion for tumor and normal tissue was similar at 3 days post cryosurgery. At 7 days after cryosurgery, the lesion was smaller for both tissues, with the normal tissue lesion being much smaller than the tumor tissue lesion. A comparison of experimental injury data to the thermal model validated in a companion paper (Hoffmann and Bischof 2001) suggested that the minimum temperature required for causing necrosis was -15.6 +/- 4.3 degrees C in tumor tissue and -19.0 +/- 4.4 degrees C in normal tissue. The other thermal parameters manifested at the edge of the lesion included a cooling rate of approximately 28 degrees C/min, 0 hold time, and a approximately 9 degrees C/min thawing rate. The conditions at the edge of the lesion are much less severe than the thermal conditions required for direct cellular destruction of AT-1 cells and tissues in vitro. These results are consistent with the hypothesis that vascular-mediated injury is responsible for the majority of injury at the edge of the frozen region in microvascular perfused tissue.     

Comparison of dual- and triple-freeze protocols for hepatic cryoablation in a Tibet pig model

The purpose of this study was to compare a dual-freeze protocol with a triple-freeze protocol for hepatic cryoablation in a porcine model. Eighteen cryoablations were performed over an exposed operation field in nine normal porcine livers, using dual- (10-5-10-5) and triple-freeze (5-5-5-5-10-5) protocols. Changes in the temperature of the cryoprobes and the diameter of the iceballs were recorded during the ablation, and pathological changes in the cryozones (zones of tissue destruction) were assessed seven days after the procedure. Use of two and three freeze-thaw cycles produced iceballs of different diameters. Seven days after cryosurgery, the triple-freeze protocol was associated with a larger zone of complete necrosis than the dual-freeze protocol, although the two protocols produced cryozones and cryolesions of similar length, and in both cases the cryozones contained five areas of destruction. With the same freezing time (20 min), the triple-freeze protocol may be a more powerful liver ablation method than the dual-freeze protocol.     

Morphologic and functional effects of renal cryoinjury

Wilms' tumor presents with bilateral renal involvement in 10% of patients. With the current treatment protocols for Wilms' tumor emphasizing maximal preservation of renal parenchyma, cryosurgery may offer an alternative to bilateral tumor resections in these patients. This study was designed to examine the effects of profound cryotherapy on a significant portion of the renal parenchyma, simulating the clinical application of this technique. Eighteen New Zealand white rabbits underwent unilateral nephrectomy, with four serving as controls. The contralateral kidney in the experimental animals was subjected to cryoinjury with a liquid nitrogen probe. Needle thermocouples monitored surface temperature 1/2 cm from the margin of the probe tip and renal "core" temperature 1 cm within the renal parenchyma directly beneath the tip. The kidney was cooled until the "core" temperature reached -40 degrees C, and this was achieved with a probe temperature of -180 degrees C for an average of 11.8 min. All animals tolerated the cryoinjury well and were sacrificed in pairs at 24 and 72 hr, and 1, 2, 3, 4, and 8 weeks. Transient gross hematuria was noted in 25% of the animals and microscopic hematuria in 50%. Serum blood urea nitrogen and creatinine levels reached maximum levels at 72 hr (BUN, 92 +/- 28 mg%; Creatinine, 7.2 +/- 1 mg%) and gradually returned toward normal thereafter. The cryolesion at 24 hr resembled a hemorrhagic infarct involving 40% of the kidney. By 8 weeks this lesion had contracted to a fibrotic lesion involving 20% of the kidney by weight. The surrounding uninjured renal parenchyma underwent compensatory hypertrophy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cryoanalgesia: electrophysiology at different temperatures

Somatosensory evoked potentials (SEP) and sensory conduction velocity (SCV) were measured in rabbit sciatic nerves following graded cold lesioning. The SEP disappeared when injury was induced at temperatures below -60 degrees C, but returned on day 41+/-4 (mean+/-SD). SEP returned on day 56+/-11 days when the lesion was induced at 100 to -180 degrees C. The SEP latency was prolonged after creating lesions at -100 to -180 degrees C, compared with both the sham operated and the -20 degrees C groups. These experiments suggest the cryolesions produced at temperatures between -60 and -100 degrees C are most suitable for altering the electrophysiological conduction of the nerve, and may result in suitable post-operative analgesia.     

Thermal dosimetry of normal porcine tissue

The response of normal porcine fat and muscle to graduated doses of hyperthermia provided by an annularly focused acoustic source was measured. Temperatures and exposure times were varied between 43 degrees C (20-90 min), 45 and 47 degrees C (20-60 min), and 49 degrees C (20 min). Response, based on histologic grading of the treated sites 30 days after exposure, was found to correlate well when mapped against several methods of estimating thermal energy deposition. The threshold for damage production was at or near 43 degrees C. For a given temperature, a nearly exponential increase in relative tissue damage as a function of increased exposure time was found. A twofold increase in tissue damage was produced in fat relative to muscle at any given thermal dose.     

The Value of Proton Magnetic Resonance Spectroscopy in High-Intensity Focused Ultrasound Treatment of Experimental Liver Cancer

High-intensity focused ultrasound (HIFU) is a rapidly developing, non-invasive technique for local treatment of solid tumors that produce coagulative tumor necrosis. This study is aimed to investigate the feasibility of proton magnetic resonance spectroscopy (MRS) on early assessing treatment of HIFU ablation in rabbit with VX2 liver tumor. HIFU ablation was performed on normal liver and VX2 tumor in rabbit, and MRS was performed on normal liver and VX2 tumor before and 2 days after 100% HIFU ablation or 80% ablation in tumor volume. Choline (Cho) and choline/lipid (Cho/Lip) ratios between complete and partial HIFU ablation of tumor were compared. Tissues were harvested and sequentially sliced to confirm the necrosis. In normal liver, the Cho value liver was not obviously changed after HIFU (P > .05), but the Cho/Lip ratio was decreased (P < .05). Cho in liver VX2 tumor was much higher than that in normal liver (P < .001). Cho and Cho/Lip ratio were significantly decreased in tumor after complete HIFU ablation and partial HIFU ablation, and the Cho value in complete HIFU tumor ablation did not show any difference from that in normal liver after HIFU (P > .05); however, the Cho value in partial ablation was still higher than that in normal liver before or in tumor after complete HIFU treatment due to the residual part of tumors, and Cho/Lip ratio is lower than that in complete HIFU treatment (P < .001). The changes in MRS parameters were consistent with histopathologic changes of the tumor tissues after treatment. MRS could differentiate the complete tumor necrosis from residual tumor tissue, when combined with magnetic resonance imaging. We conclude that MRS may be applied as an important, non-invasive biomarker for monitoring the thoroughness of HIFU ablation.     

Erythrocyte sedimentation rate. I. Volume fraction dependence in saline solution

We have measured volume fraction dependence of the sedimentation curve of swine erythrocytes in a physiological saline solution at 10 degrees C, 20 degrees C, 30 degrees C and 40 degrees C. The sedimentation curves were found to consist of initial constant velocity region and final plateau region at the lower temperatures of 10 degrees C and 20 degrees C, while modified S-shaped curves were observed at the higher temperatures of 30 degrees C and 40 degrees C. The volume fraction dependence of the initial slope v of the sedimentation curve was fitted well to the following exponential type equation at all the temperatures: v = vs,exp (1 - H)exp[-(BH + CH2)] where vs,exp is the velocity in infinite dilution corresponding to the Stokes velocity and H is the volume fraction of erythrocytes. The volume fraction dependence of the relative velocity v/vs,exp was in close agreement with a semi-empirical equation derived for slurrys in the field of chemical engineering at the lower temperatures, while a small deviation between the observed and calculated curves was found at the higher temperatures. The volume fraction dependence of v at 20 degrees C was also analyzed on a theory recently developed by Oka. The explicit functional form of the medium up-flow factor phi (H) and the deformability factor f in the theory were determined using the experimental data.     

The role of vision and proprioception in the aversion of rats to the open arms of an elevated plus-maze

The elevated plus-maze test is usually run with a short edge surrounding the open arms in order to prevent the rats from falling. The present experiment investigated the role of transparent edges differing in heights: 1 (used as control), 5, 10, 20 and 40 cm, the latter the same height as the closed arm walls. Additionally, this 40-cm high transparent edge was also studied covered by white translucent or black opaque paper. The data show that the time spent in the open arms was significantly greater when the edge height was 5, 10 or 40 cm covered by the white or black paper. However, there were no differences from the 1-cm control edge when the height was 40 cm transparent. A similar effect was observed when entries in the open arms and total entries were analyzed. The facts that there were no differences when the open arms were surrounded by 1- or 40-cm transparent edges (which allow thigmotaxis) and that the same 40-cm edge caused increases in exploratory behavior when covered by papers indicate that vision triggers aversion to open spaces.     

Cryohemostasis of uncontrolled hemorrhage from liver injury

Uncontrolled hemorrhage is the primary cause of death in both blunt and penetrating liver trauma. Cryohemostasis was attempted in the past for elective liver surgery but did not gain popularity. During past decades, cryoequipment was refined and successfully used for tumor ablation. The purpose of the present study was to assess the efficacy of cryosurgery as a potential adjuvant hemostatic technique in the treatment of grades III-IV liver injuries. A standard liver crush-evulsion injury was created in pigs. In the control group, the liver was left to bleed freely. In the experimental group, the severed liver surface was immediately frozen to -160 degrees C for 10 min, spontaneously thawed, and left to bleed thereafter. Blood pressure, pulse rate, urine output, and serum lactate were monitored. The total blood loss was measured 180 min after liver injury was inflicted. The volume of frozen liver parenchyma was measured. For further laboratory evaluation, three additional experimental animals were not sacrificed and recovered. Cryohemostasis significantly reduced blood loss and substantially attenuated hemorrhagic shock. The frozen liver parenchyma underwent necrosis but did not jeopardize survival. Cryosurgery may be an efficient adjuvant technique in the early control of hemorrhage in grades III-IV liver injury.     

Microscopic and Calorimetric Assessment of Freezing Processes in Uterine Fibroid Tumor Tissue

The use of cryosurgery in the treatment of uterine fibroids is emerging as a possible treatment modality. The two known mechanisms of direct cell injury during the tissue freezing process are linked to intracellular ice formation and cellular dehydration. These processes have not been quantified within uterine fibroid tumor tissue. This study reports the use of a combination of freeze-substitution microscopy and differential scanning calorimetry (DSC) to quantify freeze-induced dehydration within uterine fibroid tumor tissue. Stereological analysis of histological tumor sections was used to obtain the initial cellular volume (V(o)) or the Krogh model dimensions (deltaX, the distance between the microvascular channels = 15.5 microm, r(vo), the initial radius of the extracellular space = 4.8 micro m, and L, the axial length of the Krogh cylinder = 19.1 microm), the interstitial volume ( approximately 23%), and the vascular volume ( approximately 7%) of the fibroid tumor tissue. A Boyle-van't Hoff plot was then constructed by examining freeze-substituted micrographs of "equilibrium"-cooled tissue slices to obtain the osmotically inactive cell volume, V(b) = 0.47V(o). The high interstitial volume precludes the use of freeze-substitution microscopy data to quantify freeze-induced dehydration. Therefore, a DSC technique, which does not suffer from this artifact, was used to obtain the water transport data. A model of water transport was fit to the calorimetric data at 5 and 20 degrees C/min to obtain the "combined best fit" membrane permeability parameters of the embedded fibroid tumor cells, assuming either a Krogh cylinder geometry, L(pg) = 0.92 x 10(-13) m(3)/Ns (0.55 microm/min atm) and E(Lp) = 129.3 kJ/mol (30.9 kcal/mol), or a spherical cell geometry (cell diameter = 18.3 microm), L(pg) = 0.45 x 10(-13) m(3)/Ns (0.27 microm/min atm) and E(Lp) = 110.5 kJ/mol (26.4 kcal/mol). In addition, numerical simulations were performed to generate conservative estimates, in the absence of ice nucleation between -5 and -30 degrees C, of intracellular ice volume in the tumor tissue at various cooling rates typical of those experienced during cryosurgery (< or =100 degrees C/min). With this assumption, the Krogh model simulations showed that the fibroid tumor tissue cells cooled at rates < or = 50 degrees C/min are essentially dehydrated; however, at rates >50 degrees C/min the amount of water trapped within the tissue cells increases rapidly with increasing cooling rate, suggesting the formation of intracellular ice.     

Activation parameters of the blue shift (Shibata shift) subsequent to protochlorophyllide phototransformation

The Shibata shift was analyzed in flash irradiated wheat (Triticum aestivum, L., cult. MV17) leaf homogenates in the pressure range of 0.1 to 500 MPa, at temperatures of 20, 30 and 40 degrees C. The kinetics of the blue shift (called Shibata shift in case of intact leaves) was followed by repeated recording of fluorescence emission spectra after phototransformation. At 20 degrees C, above 100 MPa, the blue shift slowed down remarkably. Two components of the blue shift could be distinguished, one was pressure-dependent and the other was almost pressure-independent. The pressure-independent component can be associated with minor conformational changes of the NADPH:protochlorophyllide oxidoreductase (POR) enzyme, followed by molecular movements of the newly formed chlorophyllide molecules. The calculated activation volume of the pressure-dependent component was 43+/-11 cm(3) mol(-1) at 20 degrees C. This value reflects major molecular reorganizations in the lipid system of the membrane and in the chlorophyllide-protein complexes, and corresponds to changes of the tertiary structure of proteins which can proceed directly or indirectly via structural changes of the membrane lipids. The process was inhibited by 300 and 400 MPa at 30 and 40 degrees C, respectively. The activation volume reduced to 35+/-1.5 cm(3) mol(-1) at 40 degrees C. The decrease of the activation volume with increasing temperature indicates that the blue shift requires loosened lipid structures. The activation energy of the blue shift (measured between 10 and 40 degrees C at atmospheric pressure) was 100+/-20 kJ/mol, indicating that the structural change involves rearrangement of strong molecular interactions.     

Mitochondrial respiration in muscle and liver from cold-acclimated hypothyroid rats.

The effects of long-term cold exposure on muscle and liver mitochondrial oxygen consumption in hypothyroid and normal rats were examined. Thyroid ablation was performed after 8-wk acclimation to 4 degrees C. Hypothyroid and normal controls remained in the cold for an additional 8 wk. At the end of 16-wk cold exposure, all hypothyroid rats were alive and normothermic and had normal body weight. At ambient temperature (24 degrees C), thyroid ablation induced a 65% fall in muscle mitochondrial oxygen consumption, which was reversed by thyroxine but not by norepinephrine administration. After cold acclimation was reached, suppression of thyroid function reduced muscle mitochondrial respiration by 30%, but the hypothyroid values remained about threefold higher than those in hypothyroid muscle in the warm. Blockade of beta- and alpha1-adrenergic receptors in both hypothyroid and normal rats produced hypothermia in vivo and a fall in muscle, liver, and brown adipose tissue mitochondria respiration in vitro. In normal rats, cold acclimation enhanced muscle respiration by 35%, in liver 18%, and in brown adipose tissue 450% over values in the warm. The results demonstrate that thyroid hormones, in the presence of norepinephrine, are major determinants of thermogenic activity in muscle and liver of cold-acclimated rats. After thyroid ablation, cold-induced nonshivering thermogenesis replaced 3,5,3'-triiodothyronine-induced thermogenesis, and normal body temperature was maintained.     

Endometrial thermal balloon ablation using a high temperature, pulsed system: a mathematical model

A new endometrial thermal balloon ablation treatment for menorrhagia is modeled mathematically to predict its efficacy and safety. A device preheats a fluid to 173 degrees C within a reservoir external to the uterus, and then pulses this fluid without further heating between the reservoir and the balloon for 2.1 min of treatment time. The model predicted this treatment to result in consistent immediate tissue death (coagulation) depths of 3.4 +/- 0.1 mm for uterine cavities of 7 to 26 mL, and that eventual necrosis (tissue death that would occur 1-5 days post burn) may occur to depths of 6.5 +/- 0.2 mm. Whereas, burn depths varied with uterine cavity volume when a low temperature treatment (constant 75 degrees C for 15 min) was modeled (2.3-2.9 mm and 6.8-8.2 mm, for immediate tissue death and eventual necrosis respectively). Similarly, the high temperature, pulsed treatment was less sensitive to blood perfusion rate than the low temperature treatment. Predicted eventual necrosis depth was 1.5 mm less for the high temperature, pulsed treatment than that predicted for a low temperature treatment (constant 87 degrees C for 7 min) for the same immediate tissue death depth (3.5 mm), indicating that the new high temperature treatment may result in less damage to non targeted tissues.