Effects of capillary red cell density on gas conductance of frog skin.

We tested experimentally the hypothesis that decreasing capillary red blood cell (RBC) density (dRBC) reduces the tissue diffusing capacity of frog skin to CO (DtiCO) and O2 (DtiO2). The effects of dRBC on CO2 transport were also assessed. C18O, O2, and CO2 transport between the skin and a cutaneous sample chamber on the belly of anesthetized (halothane) frogs (Rana pipiens) was measured by mass spectrometry, and the cutaneous conductances to C18O (GCO), O2 (GO2), and CO2 (GCO2) were calculated. The dRBC of the planar cutaneous capillary bed was measured by intravital fluorescent video microscopy. DtiCO and DtiO2 were calculated from a modification of the Roughton-Foster equation: 1/G = 1/Dti + 1/(theta RBC.dRBC), where theta RBC values were estimated from literature values. In one group of animals (n = 6), measurements were made before hemodilution (dRBC = 630 +/- 56 cells/mm2), after one hemodilution (dRBC = 349 +/- 50 cells/mm2), and after a second hemodilution (dRBC = 150 +/- 31 cells/mm2). In controls, time had no effect on GCO, GO2, or GCO2 (P greater than 0.42). Before hemodilution, GCO, GO2, and GCO2 were 0.069 +/- 0.010, 0.088 +/- 0.0012, and 1.23 +/- 0.010 nmol.min-1.Torr-1.cm-2, respectively, and lowering dRBC by hemodilution decreased all these parameters (P less than 0.025). The mean slopes of GCO, GO2, and GCO2 vs. dRBC were 6.0 +/- 1.3 x 10(-7), 7.2 +/- 2.3 x 10(-7), and 7.8 +/- 3.0 x 10(-6) nmol.min-1.Torr-1.RBC-1, respectively. Lowering dRBC also decreased DtiCO and DtiO2 (P less than 0.034). DtiCO and DtiO2 were 0.080 +/- 0.012 and 0.096 +/- 0.013 nmol.min-1.Torr-1.cm-2, respectively, before hemodilution. The mean slopes of DtiCO and DtiO2 vs. dRBC were 4.9 +/- 2.1 x 10(-7) and 6.5 +/- 2.8 x 10(-7) nmol.min-1.Torr-1.RBC-1, respectively. Hemodilution had no effect on perfused capillary density (P = 0.38). These results indicate that tissue diffusive conductance is proportional to dRBC. Regulation of dRBC may be an important mechanism modulating diffusive gas transport in tissue.     

华南丘陵区冬闲稻田二氧化碳、甲烷和氧化亚氮的排放特征

采用静态箱 气相色谱法对收获后冬闲稻田CO₂、CH₄和N₂O排放进行了田间原位测定,探讨了越冬稻田3种温室气体的排放规律.结果表明,残茬稻田和裸田的CO₂的排放峰值分别出现在18:00和16:00左右.日间CH₄排放为净值,夜间表现为弱吸收.残茬稻田和裸田N₂O夜间排放分别为日间平均的1.79和1.58倍.残茬稻田的昼夜CO₂平均排放通量显著高于裸田(P＜0.05).在测定期间,残茬稻田CO₂排放随温度升高而增高.相关分析表明,CO₂排放与土温、地表温度和气温均呈显著相关,表明温度是影响收获后稻田CO₂排放的主要因素.在11月10日至翌年1月18日测定期间,残茬稻田的CO₂和CH₄平均排放通量分别为(180.69±21.21) mg·m^-2·h^-1和(-0.04±0.01) mg·m^-2·h^-1,CO₂排放通量较裸田高13.06％,CH₄吸收增高50%.残茬稻田的N₂O排放通量为(21.26±19.31) μg·m^-2·h^-1,较裸田低60.75％.由此说明华南丘陵区冬闲稻田是大气CO₂和N₂O的源,CH₄的汇.     

Permeability of human medial collateral ligament in compression transverse to the collagen fiber direction

This study quantified the apparent and intrinsic hydraulic permeability of human medial collateral ligament (MCL) under direct permeation transverse to the collagen fiber direction. A custom permeation device was built to apply flow across cylindrical samples of ligament while monitoring the resulting pressure gradient. MCLs from 5 unpaired human knees were used (donor age 55 +/- 16 yr, 4 males, 1 female). Permeability measurements were performed at 3 levels of compressive pre-strain (10%, 20% and 30%) and 5 pressures (0.17, 0.34, 1.03, 1.72 and 2.76 MPa). Apparent permeability was determined from Darcy's law, while intrinsic permeability was determined from the zero-pressure crossing of the pressure-permeability curves at each compressive pre-strain. Resulting data were fit to a finite deformation constitutive law [Journal of Biomechanics 23 (1990) 1145-1156]. The apparent permeability of human MCL ranged from 0.40 +/- 0.05 to 8.60 +/- 0.77 x 10(-16) m(4)/Ns depending on pre-strain and pressure gradient. There was a significant decrease in apparent permeability with increasing compressive pre-strain (p=0.024) and pressure gradient (p<0.001), and there was a significant interaction between the effects of compressive pre-strain and pressure (p<0.001). Intrinsic permeability was 14.14 +/- 0.74, 6.30 +/- 2.13 and 4.29 +/- 1.71 x 10(-16) m(4)/Ns for compressive pre-strains of 10%, 20% and 30%, respectively. The intrinsic permeability showed a faster decrease with increasing compressive pre-strain than that of bovine articular cartilage. These data provide a baseline for investigating the effects of disease and chemical modification on the permeability of ligament and the data should also be useful for modeling the poroelastic material behavior of ligaments.     

Reconstituting the barrier properties of a water-tight epithelial membrane by design of leaflet-specific liposomes

To define aspects of lipid composition and bilayer asymmetry critical to barrier function, we examined the permeabilities of liposomes that model individual leaflets of the apical membrane of a barrier epithelium, Madin-Darby canine kidney type 1 cells. Using published lipid compositions we prepared exofacial liposomes containing phosphatidylcholine, sphingomyelin, glycosphingolipids, and cholesterol; and cytoplasmic liposomes containing phosphatidylethanolamine, phosphatidylserine, and cholesterol. The osmotic permeability of cytoplasmic liposomes to water (P(f)), solutes, and NH(3) was 18-90-fold higher than for the exofacial liposomes (P(f(ex)) = 2.4 +/- 0.4 x 10(-4) cm/s, P(f(cy)) = 4.4 +/- 0.3 x 10(-3) cm/s; P(glycerol(ex)) = 2.5 +/- 0.3 x 10(-8) cm/s, P(glycerol(cy)) = 2.2 +/- 0.02 x 10(-6) cm/s; P(NH3(ex)) = 0. 13 +/- 0.4 x 10(-4) cm/s, P(NH3(cy)) = 7.9 +/- 1.0 x 10(-3) cm/s). By contrast, the apparent proton permeability of exofacial liposomes was 4-fold higher than cytoplasmic liposomes (P(H+(ex)) = 1.1 +/- 0. 1 x 10(-2) cm/s, P(H+(cy)) = 2.7 +/- 0.6 x 10(-3) cm/s). By adding single leaflet permeabilities, we calculated a theoretical P(f) for a Madin-Darby canine kidney apical membrane of 4.6 x 10(-4) cm/s, which compares favorably with experimentally determined values. In exofacial liposomes lacking glycosphingolipids or sphingomyelin, permeabilities were 2-7-fold higher, indicating that both species play a role in barrier function. Removal of cholesterol resulted in 40-280-fold increases in permeability. We conclude: 1) that we have reconstituted the biophysical properties of a barrier membrane, 2) that the barrier resides in the exofacial leaflet, 3) that both sphingomyelin and glycosphingolipids play a role in reducing membrane permeability but that there is an absolute requirement for cholesterol to mediate this effect, 4) that these results further validate the hypothesis that each leaflet offers an independent resistance to permeation, and 5) that proton permeation was enhanced by sphingolipid/cholesterol interactions.     

Comparative study of the O(2), CO(2) and temperature effect on respiration between "Conference" pear cell protoplasts in suspension and intact pears

The influence of the O(2) and CO(2) concentration and the temperature on the O(2) uptake rate of cool-stored intact pears and pear cell protoplasts in suspension was compared. Protocols to isolate pear cell protoplasts from pear tissue and two methods to measure protoplast respiration have been developed. Modified Michaelis-Menten kinetics were applied to describe the effect of the O(2) and the CO(2) concentration on the O(2) uptake rate and temperature dependence was analysed with an Arrhenius equation. Both systems were described with a non-competitive type of CO(2) inhibition. Due to the inclusion of gas diffusion properties, the Michaelis-Menten constant for intact pears (2.5 mM) was significantly larger than the one for protoplasts in suspension (3 microM), which was in turn larger than the Michaelis-Menten constant obtained in mitochondrial respiration measurements described in the literature. It was calculated that only 3.6% of the total diffusion effect absorbed in the Michaelis-Menten constant for intact pears, could be attributed to intracellular gas diffusion. The number of cells per volume of tissue was counted microscopically to establish a relationship between the pear cell protoplast and intact pear O(2) uptake rate. A remarkable similarity was observed: values of 61.8 nmol kg(-1) s(-1) for protoplasts and 87.1 nmol kg(-1) s(-1) for intact pears were obtained. Also, the inhibitory effect of CO(2) on the respiration rate was almost identical for protoplasts and intact pears, suggesting that protoplast suspensions are useful for the study of other aspects of the respiration metabolism.     

Peroxynitrite scavenging by ferrous truncated hemoglobin GlbO from Mycobacterium leprae

Mycobacterium leprae GlbO has been proposed to represent merging of both O(2) uptake/transport and scavenging of nitrogen reactive species. Peroxynitrite reacts with M. leprae GlbO(II)-NO leading to GlbO(III) via the GlbO(III)-NO species. The value of the second order rate constant for GlbO(III)-NO formation is >1x10(8)M(-1)s(-1) in the absence and presence of CO(2) (1.2x10(-3)M). The CO(2)-independent value of the first order rate constant for GlbO(III)-NO denitrosylation is (2.5+/-0.4)x10(1)s(-1). Furthermore, peroxynitrite reacts with GlbO(II)-O(2) leading to GlbO(III) via the GlbO(IV)O species. Values of the second order rate constant for GlbO(IV)O formation are (4.8+/-0.5)x10(4) and (6.3+/-0.7)x10(5)M(-1)s(-1) in the absence and presence of CO(2) (=1.2x10(-3)M), respectively. The value of the second order rate constant for the peroxynitrite-mediated GlbO(IV)O reduction (= (1.5+/-0.2)x10(4)M(-1)s(-1)) is CO(2)-independent. These data argue for a role of GlbO in the defense of M. leprae against nitrosative stress.     

Transport phenomena during freezing of adipose tissue derived adult stem cells

In the present study a well-established differential scanning calorimeter (DSC) technique is used to measure the water transport phenomena during freezing of stromal vascular fraction (SVF) and adipose tissue derived adult stem (ADAS) cells at different passages (Passages 0 and 2). Volumetric shrinkage during freezing of adipose derived cells was obtained at a cooling rate of 20 degrees C/min in the presence of extracellular ice and two different, commonly used, cryoprotective agents, CPAs (10% DMSO and 10% Glycerol). The adipose derived cells were modeled as spheres of 50 microm diameter with an osmotically inactive volume (Vb) of 0.6Vo, where Vo is the isotonic cell volume. By fitting a model of water transport to the experimentally obtained volumetric shrinkage data, the "best-fit" membrane permeability parameters (reference membrane permeability to water, Lpg or Lpg[cpa] and the activation energy, ELp or ELp[cpa]) were determined. The "best-fit" membrane permeability parameters for adipose derived cells in the absence and presence of CPAs ranged from: Lpg=23.1-111.5x10(-15) m3/Ns (0.135-0.652 microm/min-atm) and ELp=43.1-168.8 kJ/mol (9.7-40.4 kcal/mol). Numerical simulations of water transport were then performed under a variety of cooling rates (5-100 degrees C/min) using the experimentally determined membrane permeability parameters. And finally, the simulation results were analyzed to predict the optimal rates of freezing adipose derived cells in the presence and absence of CPAs.     

Water permeability of asymmetric planar lipid bilayers: leaflets of different composition offer independent and additive resistances to permeation

To understand how plasma membranes may limit water flux, we have modeled the apical membrane of MDCK type 1 cells. Previous experiments demonstrated that liposomes designed to mimic the inner and outer leaflet of this membrane exhibited 18-fold lower water permeation for outer leaflet lipids than inner leaflet lipids (Hill, W.G., and M.L. Zeidel. 2000. J. Biol. Chem. 275:30176-30185), confirming that the outer leaflet is the primary barrier to permeation. If leaflets in a bilayer resist permeation independently, the following equation estimates single leaflet permeabilities: 1/P(AB) = 1/P(A) + 1/P(B) (Eq. l), where P(AB) is the permeability of a bilayer composed of leaflets A and B, P(A) is the permeability of leaflet A, and P(B) is the permeability of leaflet B. Using for the MDCK leaflet-specific liposomes gives an estimated value for the osmotic water permeability (P(f)) of 4.6 x 10(-4) cm/s (at 25 degrees C) that correlated well with experimentally measured values in intact cells. We have now constructed both symmetric and asymmetric planar lipid bilayers that model the MDCK apical membrane. Water permeability across these bilayers was monitored in the immediate membrane vicinity using a Na+-sensitive scanning microelectrode and an osmotic gradient induced by addition of urea. The near-membrane concentration distribution of solute was used to calculate the velocity of water flow (Pohl, P., S.M. Saparov, and Y.N. Antonenko. 1997. Biophys. J. 72:1711-1718). At 36 degrees C, P(f) was 3.44 +/- 0.35 x 10(-3) cm/s for symmetrical inner leaflet membranes and 3.40 +/- 0.34 x 10(-4) cm/s for symmetrical exofacial membranes. From, the estimated permeability of an asymmetric membrane is 6.2 x 10(-4) cm/s. Water permeability measured for the asymmetric planar bilayer was 6.7 +/- 0.7 x 10(-4) cm/s, which is within 10% of the calculated value. Direct experimental measurement of P(f) for an asymmetric planar membrane confirms that leaflets in a bilayer offer independent and additive resistances to water permeation and validates the use of.     

Predicted permeability parameters of human ovarian tissue cells to various cryoprotectants and water

This study presents a generic numerical model to simulate the coupled solute and solvent transport in human ovarian tissue sections during addition and removal of chemical additives or cryoprotective agents (CPA). The model accounts for the axial and radial diffusion of the solute (CPA) as well as axial convection of the CPA, and a variable vascular surface area (A) during the transport process. In addition, the model also accounts for the radial movement of the solvent (water) into and out of the vascular spaces. Osmotic responses of various cells within an human ovarian tissue section are predicted by the numerical model with three model parameters: permeability of the tissue cell membrane to water (L(p)), permeability of the tissue cell membrane to the solute or CPA (omega) and the diffusion coefficient of the solute or CPA in the vascular space (D). By fitting the model results with published experimental data on solute/water concentrations within an human ovarian tissue section, I was able to determine the permeability parameters of ovarian tissue cells in the presence of 1.5M solutions of each of the following: dimethyl sulphoxide (DMSO), propylene glycol (PROH), ethylene glycol (EG), and glycerol (GLY), at two temperatures (4 degrees C and 27 degrees C). Modeling Approach 1: Assuming a constant value of solute diffusivity (D = 1.0 x 10(-9) m(2)/sec), the best fit values of L(p) ranged from 0.35 x 10(-14) to 1.43 x 10(-14) m(3)/N-sec while omega ranged from 2.57 x 10(-14) to 70.5 x 10(-14) mol/N-sec. Based on these values of L(p) and omega, the solute reflection coefficient, sigma defined as sigma = 1-omega v(CPA)/L(P) ranged from 0.9961 to 0.9996. Modeling Approach 2: The relative values of omega and sigma from our initial modeling suggest that the embedded ovarian tissue cells are relatively impermeable to all the CPAs investigated (or omega approximately 0 and sigma approximately 1.0). Consequently the model was modified and used to predict the values of L(p) and D assuming omega = 0 and sigma = 1.0. The best fit values of L(p) ranged from 0.44 x 10(-14) to 1.2 x 10(-14) m(3)/N-sec while D ranged from 0.85 x 10(-9) to 2.08 x 10(-9) m(2)/sec. Modeling Approach 3: Finally, the best fit values of D from modeling approach 2 were incorporated into model 1 to re-predict the values of L(p) and omega. It is hoped that the ovarian tissue cell parameters reported here will help to optimize chemical loading and unloading procedures for whole ovarian tissue sections and consequently, tissue cryopreservation procedures.     

Reductive nitrosylation and peroxynitrite-mediated oxidation of heme-hemopexin

Hemopexin (HPX), which serves as a scavenger and transporter of toxic plasma heme, has been postulated to play a key role in the homeostasis of NO. In fact, HPX-heme(II) reversibly binds NO and facilitates NO scavenging by O(2). HPX-heme is formed by two four-bladed beta-propeller domains. The heme is bound between the two beta-propeller domains, residues His213 and His266 coordinate the heme iron atom. HPX-heme displays structural features of heme-proteins endowed with (pseudo-)enzymatic activities. In this study, the kinetics of rabbit HPX-heme(III) reductive nitrosylation and peroxynitrite-mediated oxidation of HPX-heme(II)-NO are reported. In the presence of excess NO, HPX-heme(III) is converted to HPX-heme(II)-NO by reductive nitrosylation. The second-order rate constant for HPX-heme(III) reductive nitrosylation is (1.3 +/- 0.1) x 10(1) m(-1).s(-1), at pH 7.0 and 10.0 degrees C. NO binding to HPX-heme(III) is rate limiting. In the absence and presence of CO2 (1.2 x 10(-3) m), excess peroxynitrite reacts with HPX-heme(II)-NO (2.6 x 10(-6) m) leading to HPX-heme(III) and NO, via the transient HPX-heme(III)-NO species. Values of the second-order rate constant for HPX-heme(III)-NO formation are (8.6 +/- 0.8) x 10(4) and (1.2 +/- 0.2) x 10(6) m(-1).s(-1) in the absence and presence of CO2, respectively, at pH 7.0 and 10.0 degrees C. The CO2-independent value of the first-order rate constant for HPX-heme(III)-NO denitrosylation is (4.3 +/- 0.4) x 10(-1) s(-1), at pH 7.0 and 10.0 degrees C. HPX-heme(III)-NO denitrosylation is rate limiting. HPX-heme(II)-NO appears to act as an efficient scavenger of peroxynitrite and of strong oxidants and nitrating species following the reaction of peroxynitrite with CO2 (e.g. ONOOC(O)O-, CO3-, and NO2).     

Diffusive and convective transport through hollow fiber membranes for liver cell culture

For an efficient membrane bioreactor design, transport phenomena determining the overall mass flux of metabolites, catabolites, cell regulatory factors, and immune-related soluble factors, need to be clarified both experimentally and theoretically. In this work, experiments and calculations aimed at discerning the simultaneous influence of both diffusive and convective mechanisms to the transport of metabolites. In particular, the transmembrane mass flux of glucose, bovine serum albumin (BSA), APO-transferrin, immunoglobulin G, and ammonia was experimentally measured, under pressure and concentration gradients, through high-flux microporous hydrophilic poly-ether-sulphone (PES-HFMs) and poly-sulphone hollow fiber membranes (PS-HFMs). These data were analyzed by means of a model based on the mechanism of capillary pore diffusion, assuming that solute spherical molecules pass through an array of solvent-filled cylindrical pores with a diffusive permeation corrected for friction and steric hindrances. Additionally, resistances to the mass transfer were taken into account. Convective permeation data were discussed in terms of morphological properties of the polymeric membranes, molecular Stokes radius, and solute-membrane interactions according to information given by contact angle measurements. The observed steady-state hydraulic permeance of PS-HFMs was 0.972 L/m2hmbar, about 15.6-fold lower than that measured for PES-HFMs (15.2 L/m2h); in general, PS-HFMs provided a significant hindrance to the transport of target species. Diffusion coefficients of metabolites were found to be similar to the corresponding values in water through PES-HFMs, but significantly reduced through PS-HFMs (D(Glucose)(Membrane)=2.8x10(-6)+/-0.6x10(-6)cm2/s, D(BSA)(Membrane)=6.4 x 10(-7)+/-1 x 10(-7)cm(/s, D(Apotransferrin)(Membrane)=2.3 x 10(-7)+/-0.25 x 10(-7)cm2/s).     

Effect of CO2 concentrations on acoustic inferences of airway area

To determine the effect of gas composition on the accuracy of measurements of airway area and distance using an acoustic reflection technique, we employed glass-tube models to simulate pharyngeal (Phar-model), laryngeal (Lar-model), and tracheal (Trach-model) regions of upper and central airways. We made repeated measurements of area-distance functions using gas mixtures containing 0, 2, 4, 6, 8, and 10% CO2, 80% He, and balance O2. The actual area of the model was calculated from the roentgenographic data and compared favorably with an area measured by acoustic reflections using a gas mixture containing 0% CO2. With the different gas mixtures, calculated area was overestimated only at the highest levels of CO2, with Phar-model area increasing from (mean +/- SD) 4.66 +/- 0.03 cm2 measured with 0% CO2 to 4.93 +/- 0.05 cm2 (P less than 0.05) measured with CO2 concentration of 10%. To assess the effect of CO2 concentration on measurements of distance, we isolated two discrete points located in the Phar-model and Lar-model regions. When measurements were performed using 10% CO2 mixture, Phar-model point was shifted by 1.02 +/- 0.03 cm and Lar-model point was shifted by 2.16 +/- 0.09 cm away from the microphone compared with their axial position determined, using 0% CO2 mixture (P less than 0.05). Differences in area-distance calculations at the higher levels of CO2 did not exceed the within-run variability of the technique (10 +/- 4%). We conclude that CO2 absorbers are not required during measurements of airway area by acoustic reflections, provided CO2 concentration does not exceed 10%.     

Experimental and theoretical investigation of directional permeability of human vertebral cancellous bone for cement infiltration

The use of acrylic polymers in infiltrating the porous bone structure is an emerging procedure for the augmentation of osteoporotic vertebrae. Although this procedure is employed frequently, it is performed based on empirical knowledge, and therefore, does not take into consideration the porosity-dependent permeability of human vertebral cancellous bone. The purpose of this study was to: (a). experimentally and theoretically investigate interdependence of the vertebral cancellous bone permeability and porosity, and (b). examine if the bone permeability of spinal cancellous bone can be predicted using bone mineral density measurements. If these relations can be established, they can be useful in optimizing the injection conditions for predicable cement infiltration. To determine the porosity-dependent and directional permeability, 34 bone cores-20 samples in the superior-inferior (SI) direction and 14 in the anterior-posterior (AP) direction-were cut from 20 lumbar vertebrae and infiltrated with silicone oil with a viscosity matching that of PMMA. The permeability of the cores was determined based on Darcy's law. The mean permeability of SI and AP cores was 4.45+/-1.72 x 10(-8) and 3.44+/-1.26 x 10(-8)m(2), respectively. An interesting finding of this study was that the permeability of the AP cores was approximately 78% of that of SI cores, though the porosity of the SI and AP cores taken from the same vertebra was approximately equal. In addition, we provided a theoretical model for the porosity-dependent permeability that accurately described non-linear dependency of the bone permeability and porosity in both directions. Although the relation of the bone permeability and porosity was established, bone mineral density was a weak predictor of the bone permeability. The experimental and theoretical results of this study can be used to understand polymer flow in cement infiltration procedures.     

Phosphoenolpyruvate carboxylase from cherimoya fruit: properties, kinetics and effects of high CO(2).

Phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) regulatory properties were studied in non-photosynthetic (mesocarp) and photosynthetic (peel) tissues from cherimoya (Annona cherimola Mill.) fruit stored in air, in order to gain a better understanding of in vivo enzyme regulation. Analyses were also performed with fruit treated with 20% CO(2)-20% O(2) to define the role of PEPC as part of an adaptive mechanism to high external carbon dioxide levels. The results revealed that the special kinetic characteristics of the enzyme from mesocarp--high V(max) and low sensibility to L-malate inhibition - are related to the active acid metabolism of these fruits and point to a high rate of reassimilation of respired CO(2) into keto-acids. With respect to fruit stored in air, PEPC in crude extracts from CO(2)-treated cherimoyas gave a similar V(max) (1.12+/-0.03 microkat x mg(-1) protein), a lower apparent K(m) (68+/-9 microM for PEP) and a higher I(50) of L-malate (5.95+/-0.3 mM). These kinetic values showed the increase in the affinity of this enzyme toward one of its substrate, PEP, by elevated external CO(2) concentrations. The lower K(m) value and lower sensitivity to L-malate are consistent with higher in vivo carboxylation reaction efficiency in CO(2)-treated cherimoyas, while pointing to an additional enzyme regulation system via CO(2).     

Effect of hypoxia on permeability of pulmonary endothelium of canine visceral pleura

To determine if hypoxia increases the permeability of the pulmonary capillaries of the visceral pleura, water and protein movement across visceral pleura of isolated blood-perfused lungs ventilated with 20% O2-5% CO2 or 0% O2-5% CO2 was analyzed in terms of a two-compartment model of fluid exchange. Lungs from mongrel dogs were enclosed in a water-impermeable membrane, thereby creating an artificial visceral pleural space (VPS); fluid flux was determined as the filtration or reabsorption of water and protein in the VPS. Hypoxic vasoconstriction was prevented by adding verapamil to the perfusate. Hydrostatic pressures were continuously monitored and samples of perfusate and pleural fluid were obtained for protein determinations. Pulmonary capillary pressure was varied between 5 and 20 Torr by changing venous pressure while the protein concentration gradient was varied from 0.5 to 6.6 g/dl by introducing different solutions of plasma mixed with saline into the VPS. The hydraulic conductivity (Lp) increased from 4.25 +/- 0.74 to 9.18 +/- 0.67 X 10(-7) ml X s-1 X mmHg-1 X cm-2 and the diffusional permeability (Pd) of protein increased from 1.29 +/- 0.28 to 4.06 +/- 0.44 X 10(-6) cm/s under hypoxic conditions (P less than 0.05). Inhibition of xanthine oxidase by the addition of allopurinol (10 mg/kg body wt) to the perfusate prevented the increase in Lp and Pd observed under hypoxic conditions. We conclude that free radicals generated via xanthine oxidase may be responsible for the increased permeability observed during severe hypoxia.     

Freezing response of white bass (Morone chrysops) sperm cells

The water transport response during freezing of sperm cells of Morone chrysops (white bass, WB) was obtained using a shape-independent differential scanning calorimeter (DSC) technique. Sperm cell suspensions were frozen at a cooling rate of 20 degrees C/min in two different media: (1) without cryoprotective agents (CPAs), or (2) with 5% (v/v) dimethyl sulfoxide (Me2SO). For calculations, the sperm cell was modeled as a cylinder of length 24.8 microm and diameter of 0.305 microm, while the osmotically inactive cell volume (Vb) was assumed to be 0.6 Vo, where Vo was the isotonic or the initial cell volume. By fitting a model of water transport to the experimentally determined water transport data, the best fit membrane permeability parameters (reference membrane permeability to water, Lpg or Lpg[cpa] and the activation energy, ELp or ELp[cpa]) were determined, and ranged from Lpg = 0.51-1.7 x 10(-15) m3/Ns (0.003-0.01 microm/min-atm), and ELp = 83.6-131.3 kJ/mol (20.0-31.4 kcal/mol). The parameters obtained in this study suggest that the optimal rate of cooling for M. chrysops sperm cells is approximately 22 degrees C/min, a value that compares closely with experimentally determined optimal rates of cooling (approximately 16 degrees C/min).     

Exploring gas permeability of cellular membranes and membrane channels with molecular dynamics

Aquaporins are a family of membrane proteins specialized in rapid water conduction across biological membranes. Whether these channels also conduct gas molecules and the physiological significance of this potential function have not been well understood. Here we report 140 ns of molecular dynamics simulations of membrane-embedded AQP1 and of a pure POPE bilayer addressing these questions. The permeability of AQP1 to two types of gas molecules, O2 and CO2, was investigated using two complementary methods, namely, explicit gas diffusion simulation and implicit ligand sampling. The simulations show that the central (tetrameric) pore of AQP1 can be readily used by either gas molecule to permeate the channel. The two approaches produced similar free energy profiles associated with gas permeation through the central pore: a -0.4 to -1.7 kcal/mol energy well in the middle, and a 3.6-4.6 kcal/mol energy barrier in the periplasmic vestibule. The barrier appears to be mainly due to a dense cluster of water molecules anchored in the periplasmic mouth of the central pore by four aspartate residues. Water pores show a very low permeability to O2, but may contribute to the overall permeation of CO2 due to its more hydrophilic nature. Although the central pore of AQP1 is found to be gas permeable, the pure POPE bilayer provides a much larger cross-sectional area, thus exhibiting a much lower free energy barrier for CO2 and O2 permeation. As such, gas conduction through AQP1 may only be physiologically relevant either in membranes of low gas permeability, or in cells where a major fraction of the cellular membrane is occupied by AQPs.     

Separate effects of ischemia and reperfusion on vascular permeability in ventilated ferret lungs.

In systemic organs, ischemia-reperfusion injury is thought to occur during reperfusion, when oxygen is reintroduced to hypoxic ischemic tissue. In contrast, the ventilated lung may be more susceptible to injury during ischemia, before reperfusion, because oxygen tension will be high during ischemia and decrease with reperfusion. To evaluate this possibility, we compared the effects of hyperoxic ischemia alone and hyperoxic ischemia with normoxic reperfusion on vascular permeability in isolated ferret lungs. Permeability was estimated by measurement of filtration coefficient (Kf) and osmotic reflection coefficient for albumin (sigma alb), using methods that did not require reperfusion to make these measurements. Kf and sigma alb in control lungs (n = 5), which were ventilated with 14% O2-5% CO2 after minimal (15 +/- 1 min) ischemia, averaged 0.033 +/- 0.004 g.min-1.mmHg-1.100 g-1 and 0.69 +/- 0.07, respectively. These values did not differ from those reported in normal in vivo lungs of other species. The effects of short (54 +/- 9 min, n = 10) and long (180 min, n = 7) ischemia were evaluated in lungs ventilated with 95% O2-5% CO2. Kf and sigma alb did not change after short ischemia (Kf = 0.051 +/- 0.006 g.min-1.mmHg-1.100 g-1, sigma alb = 0.69 +/- 0.07) but increased significantly after long ischemia (Kf = 0.233 +/- 0.049 g.min-1 x mmHg-1 x 100 g-1, sigma alb = 0.36 +/- 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)