Limitations of hairless mouse skin as a model for in vitro permeation studies through human skin: hydration damage.

Hairless mouse skin currently provides a popular model membrane for studies in human percutaneous absorption. Although some similarities between the two skin types have been demonstrated, the effects of prolonged hydration on hairless mouse skin have not previously been rigorously examined. We have measured in vitro the effects of hydration at 31 degrees C on the permeabilities of hairless mouse skin and human abdominal and scalp skin to a model polar compound (water) and a lipid material (hexanol). The permeability of hairless mouse skin rose dramatically, especially to water (fiftyfold increase), whereas the human skin was more stable. We also compared the effects of stripping the stratum corneum with the effects of 8-d hydration for hairless mouse and human abdominal skin. Hydration of hairless mouse skin was as effective as tape-stripping in eliminating the stratum corneum barrier, whereas stripping human skin was far more damaging than hydration, suggesting that prolonged hydration mechanically disrupted mouse skin but not human skin. Histological examination of fresh and hydrated tissues confirmed this suggestion. We therefore recommend that hairless mouse skin is not used as a model for human tissue during in vitro permeation studies under conditions of long-term hydration, i.e., greater than three days.

Shed snake skin and hairless mouse skin as model membranes for human skin during permeation studies.

Difficulties in obtaining and using human skin have tempted many workers to employ animal membranes for percutaneous absorption studies. We have investigated the suitability of two species of snake (Elaphe obsoleta, Python molurus) for this purpose and compared our in vitro experimental results for human skin and for hairless mouse, a currently popular model. The effects of long-term hydration on the membranes were investigated over 8 d using tritiated water as a model permeant. The initial permeability coefficients of all the membranes were similar (0.74-2.2 X 10(-3) cm 2h-1). Although the human and squamate skins did not change significantly over the test period, the permeability of hairless mouse skin increased 37 times. The actions of typical enhancers on the permeabilities of the membranes to a model penetrant 5-fluorouracil (5-FU) were tested using 3% Azone in Tween 20/saline, propylene glycol (PG), 2% Azone in PG, and 5% oleic acid in PG. While the data from snake membranes tended to underestimate the enhancer effects, those from hairless mouse skin greatly overestimated the changes. None of the membranes was a completely reliable model for assessing human percutaneous absorption as modified by accelerants. Pretreatment with acetone did not significantly change the permeability of human or squamate skins to 5-FU, although that of hairless mouse increased twentyfold. An overall conclusion is that, wherever possible, human skin should be used in absorption studies and not hairless mouse or snake skin; otherwise, misleading results may be obtained.

Evaluation of regional differences of tracer appearance time in cerebral tissues using [15O] water and dynamic positron emission tomography.

The tracer appearance time relative to the radial artery-sampling site has been evaluated in six brain locations in five human subjects using dynamic positron emission tomography (PET) following the bolus injection of H2(15)O. There was a maximum difference of +/- 2 s from the average in each location. To globally adjust the timing difference between the measured arterial curve and the PET scan, a correction method was developed based on a nonlinear least-squares fitting procedure. This new technique determined the global time delay with an accuracy of +/- 0.5 s. On the other hand, the linear backward extrapolation method resulted in a systematic error of 4 s.

Fluid transport across human fetal membranes affected by human amniotic fluid prolactin: an in vitro study.

The aim of the study was to investigate the effects of human amniotic fluid (AF) prolactin (PRL) on the transfer of fluids across human fetal membranes, detected in vitro by weight change. We have developed a system, consisting of an inner compartment (4 ml), suspended in an outer compartment (40 ml), and separated from it by a 2 cm2 circular piece of fresh human fetal membrane (amnion, chorion and adhering decidua). The weight of the inner compartment was continuously registered with an electronic precision balance. Osmolality changes or hydrostatic pressure differences did not affect the rate of mass transfer through the membranes. Potassium cyanide significantly influenced the mass transfer in favour of the applied hydrostatic pressure difference (+2 cm H2O to the amniotic side) or osmotic pressure difference (amniotic side 265 mosm/kg/decidual side 285 mosm/kg). The PRL preparations used were human and ovine pituitary PRL, as well as three fractions isolated from human AF by fractionated ammonium sulphate and ethanol precipitations, followed by Sephacryl chromatography. All PRL preparations were tested in physiologic concentrations (0.5 to 5 micrograms/ml). Only the two largest AF-PRL variants significantly disturbed the balance of fluid transfer across the membranes when added to the fetal (amniotic) side of the membrane. This resulted in a net increase in fetomaternal transfer of 120 to 180 microliters. This effect could be repeated and lasted for at least 25 minutes. Using an antibody against hPRL the effect was completely blocked. Neither AF-PRL added to the maternal (decidual) side of the membrane nor oPRL or human pituitary PRL added to the amniotic or decidual side changed the rate of mass transfer across the membranes.

Effect of low mole fraction of trehalose dicorynomycolate from Corynebacterium diphtheriae on water permeability and electrical capacitance of lipid bilayer membranes.

The effect of incorporation of different proportions of trehalose dicorynomycolates (TDC) into lecithin bilayer membranes was studied. It was found that TDC, induces a 14% decrease of water osmotic permeability (42.6 +/- 3.9 to 36.8 +/- 2.7 microns/s) at 1.6 mole%, suggesting that this substance leads to an increase of the degree of packing of the constituent lipid molecules. A condensing effect of TDC was also apparent from membrane electrical capacitance (Cm) measurement. By incorporating TDC into bilayer membranes, the value of Cm experienced a decrease of 29% at 1.6% mole fraction. This finding was taken to reflect an increase in membrane thickness, known in many examples, to be related to the condensing effect.

Analysis of intestinal water absorption and changes in circulating blood volume in rats.

To analyze the change in blood volume after intestinal absorption, circulating blood volume was continuously monitored in rats after infusion of various solutions into the small intestine. Arterial and venous catheters were connected to a system for continuous monitoring of blood volume by the dilution method using 51Cr-labeled erythrocytes. Test solutions (tap water, 0.45, 0.9, 1.8% NaCl, 0.45% NaCl with 2% glucose, and 5% glucose) were infused at a rate of 1 ml/100 g body wt. for 10 min through a duodenal catheter. After the infusion, blood volume increased except in the 1.8% NaCl group, which showed a transient decrease in blood volume by about 10%. The rate of blood volume increase was highest in the 0.45% NaCl with 2% glucose group and lowest in the 1.8% NaCl group. The retention ratio of infused solution in the vascular space was almost identical among the groups and was about 22%. These results indicate that the rate of increase in blood volume after intestinal administration of fluid is modified by the osmolality of the fluid and Na-glucose co-transport, whereas the retention ratio of the infused fluid in blood is constant.

Thermodynamic network modelling of transfer across the perfused guinea-pig placenta, using SPICE.

The response of the dually perfused guinea pig placenta to bolus inputs of the marker substances 125I-albumin and tritiated water is used to model vascular placental topology and membrane characteristics. This is accomplished by generating a thermodynamic network containing ten adjustable variables describing flow and transfer quantitatively. After translation of the network into an electronic circuit the parameter values are determined with the use of the SPICE program. The results are then used to estimate unknown physiological variables like shunted flow, membrane permeabilities, etc. This model has potential usefulness in the study of those areas of placental physiology in which transient analysis is required.

[Estimation of absorbed dose of beta radiation into the critical tissues by a single injection of tritiated water].

The biological effects of tritium in humans need to be clarified, because the chances of humans becoming exposed to tritium beta radiation may increase with the development of the nuclear fusion reactor. To evaluate the biological effects of tritium, it is necessary to estimate exactly the absorbed dose from the tritium beta rays in the tissue. In many reports, the absorbed dose of HTO in the tissues is estimated from the tritium content in body fluid and dose calculations are customarily based upon the water content of soft tissues, which is taken to be 0.7 to 0.8. However, these methods may not show the exact absorbed dose in the organs. In the present study, the radioactivity of the critical tissues was measured directly using a sample oxidizer and the absorbed dose was calculated from the radioactivity of tritium in the tissues. Details on the method for calculation of the absorbed dose in tissues of the mouse is shown in this report. The results suggest that the absorbed dose should be obtained from the radioactivity in the tissues.

On mechanisms of water nuclear magnetic relaxation in protein solutions.

To describe water nuclear magnetic relaxation in protein solutions a new modification of two-state model has been advanced. It is assumed that the hydration water molecules take part in two type rotation motions: fast anisotropic and slow isotropic ones, each of them can be characterized by a single correlation time. A transition of water molecule from bound state into bulk water is considered as defect arousing and is described by defect diffusion model. The model advanced allows one to describe both frequency and temperature dependences of water spin-lattice relaxation times in protein solutions and to get an information on microdynamic parameters of hydration water molecules such as life time, fraction, hindrance of local motion, etc.

Hydration forces involved in cellular interactions.

Cellular physical interactions are determinant in many and various biological processes: aggregation, adhesion, segregation, contact inhibition, fusion, etc. A real advance concerning the understanding of cellular interactions from a physical point of view has been performed of late, but a great many problems still remain to be clarified. Thus, DLVO theory extrapolated to cell-cell interactions, is unable to describe cellular interactions at very short distances of separation, where other forces play an important role. A sketch of the original form of DLVO theory, its criticism along with the nature and mechanism of the short range repulsive forces (hydration forces) are presented and discussed.

Permeation of water through the polymeric components of boil-in-bag laminates.

Diffusion coefficients and equilibrium uptake have been measured for water and 3% acetic acid in film of PETP, lldpe, nylon 6 and nylon 66, and also in some isocyanate adhesives. These materials are used in the construction of boil-in-bags for food. The three methods of measurement used were permeation from a bag, mass uptake, and the use of a novel technique using isotopically labelled water. The principal temperature of investigation has been 100 degrees C. Differences in the permeation parameters for water and 3% acetic acid were small for PETP, but surprisingly large for an inert material such as lldpe. With the nylons differences could readily be ascribed to the fact that these materials are bases. When immersed in boiling water there was no change in the degree of crystallinity of PETP, but lldpe and nylon 66 showed small increases.

[The effect of external gamma-irradiation on the distribution of deuterium oxide in the rat body]. / Vliianie vneshnego gamma-izlucheniia na raspredelenie okisi deiteriia v organizme krys.

External gamma-irradiation (7.74 and 15.48 x 10(-2) C/kg) does not influence the distribution of deuterium oxide within a rat body: it is distributed uniformly among the organs and tissues as it is observed in nonirradiated animals. The effect of external irradiation favors the retention of deuterium oxide within the organs and tissues. The delayed excretion of deuterium oxide from the body can enhance the biological effect. This should be taken into account in standardizing the combined radiation effects.

Root parenchyma cells in water transport.

In experiments with the "sleeves" and the whole roots of Zea mays seedings, the idea of the complicated nature of root pressure and an active part of parenchyma cells in water pumping by roots has been confirmed. It has been shown that root pumping activity is summarized by two, principally different constituents--metabolic and osmotic. The metabolic constituent functions exclusively at the expense of the parenchyma cells activity.

Definitions of differences and changes in peritoneal membrane water transport properties.

A survey is given comparing measurements of transperitoneal water transport in different clinical situations with analyses based on the so-called "pore theory." This model links the measured changes to physical alterations of the peritoneal membrane. The calculations include "equivalent pore radius," effective "membrane area" and diffusive length, the transport resistance of the unstirred dialysate layer, and the residual intraperitoneal volume after dialysate drainage. The clinical appearances include individual differences in transperitoneal transport characteristics, changes in transperitoneal transport over time on continuous ambulatory peritoneal dialysis (CAPD) and during peritonitis, the pharmacological effect on the transport properties, and the effect of peritoneal catheter dislocation on ultrafiltration capacity. The main conclusions are as follow: During CAPD treatment the measurement of intraperitoneal solute equilibration and "mass-transfer-area coefficients" for urea and creatinine is less sensitive than the measurement of ultrafiltration volume in revealing peritoneal membrane changes. Differences and changes found have mostly a combined physical explanation, but one is more or less dominant. Changes in peritoneal membrane area seem to be the most dominant cause of changes in transperitoneal transport during time on CAPD and when sodium nitroprusside was added to the peritoneal dialysate. Changes during peritonitis can be explained by changes in pore radius and depth. Individual differences can be explained by differences in "membrane" area and in resistance of the unstirred dialysate fluid. High residual dialysate volume can give rise to clinical problems and should be considered when placing the catheter in the peritoneal cavity.

Comparison of transcellular and transepithelial water osmotic permeabilities (Pos) in the isolated proximal straight tubule (PST) of the rabbit kidney.

Measurements of the water osmotic permeabilities of apical and basolateral membranes of PST cells and of the transepithelial permeability have been carried out using a very fast method with high temporal and spatial resolution. At 25 degrees C the values obtained are: 80.8 +/- 11.9 x 10(-4) cm3/s osmol cm2 of apical (luminal) surface area and 90.1 +/- 13.0 x 10(-4) cm3/s osmol cm2 of basement membrane area (no membrane invaginations taken in account). These values are higher than previously published values due to the use of a faster and more accurate volume measuring and recording system. The transepithelial water osmotic permeability at 25 degrees C is 77 +/- 11 in units of 10(-4) cm3/s osmol cm2 basement membrane area. The transcellular water osmotic permeability is 32 +/- 7 (same units), leaving a paracellular contribution of 45 +/- 10 (same units). In the presence of 2.5 mM parachloromercuribenzenesulfonate (pCMBS) the apical permeability is reduced with an incubation of 10-15 min to 23% of its control value and the basolateral permeability to 8% of its control value (after 25 min) but the transepithelial permeability is only reduced to about 1/2 of the control value. This leaves a transcellular permeability of 6 x 10(-4) cm3/s osmol cm2 of basement membrane area and a paracellular contribution of 33 +/- 6 (same units). These results indicate a significant contribution of the paracellular pathway to the transepithelial water osmotic permeabilities in PST.

Effect of cecectomy on water and nutrient absorption of birds.

The effect of the removal of the avian digestive ceca on osmoregulation and on absorption of certain nutrients is reviewed. While data indicate that the ceca have the potential for absorption of a significant quantity of water, several studies have demonstrated that effects of cecectomy on water intake and output are transitory and that compensatory adjustments made within 2 to 3 weeks postsurgery allow cecectomized birds to eat and gain weight normally. However, cecectomized great horned owls exposed to 27 degrees C turned over their body water 1.6 times faster than intact owls under the same conditions, suggesting that the ceca do have a vital role in water balance of thermally stressed horned owls. Cecectomy resulted in slightly lower metabolizability of food, lower digestibility of crude fiber in low fiber diets, and greater loss of certain amino acids in cecectomized birds that were fasted or fed a protein-free diet. The latter suggests that the ceca are important in recovery of amino acids endogenous protein degradation. Loss of cecal function also resulted in higher energy intake and excretion in Japanese quail, amounting to 5.7% of the total daily energy requirement.

Effect of chronic vanadate ingestion on amino acid and water absorption in rat intestine.

Chronic administration of a relatively low concentration of vanadate to rats causes inhibition of water, alpha-aminoisobutyric acid (AIB) and L-alanine absorption. The mechanism responsible for this inhibition was tested by studying the uptake of alanine in isolated rat intestinal cells. The studies suggest that the vanadate inhibition of amino acid transport is primarily caused by a decreased activity of the Na+-K+ pump, an action that is similar to what is observed when the rat intestine is acutely exposed to vanadate. Vanadate appeared to have no direct effect on the entry of amino acids into the intestinal cell. This was evident by the fact that amino acid uptake by enterocytes of control rats was not different from the uptake by cells of vanadate-treated animals that have an inwardly directed Na gradient artificially created across them. Furthermore, 86RB influx and efflux into and out of intestinal tissues of the vanadate-treated animals were, respectively, decreased and increased as compared to normal control tissues and they were similar to what is observed when the intestine is acutely exposed to ouabain, a known specific inhibitor of the Na+-K+ pump.

Lung vascular permeability changes in lambs with hyaline membrane disease.

To investigate the mechanism of pulmonary edema in hyaline membrane disease (HMD), lymph from the efferent duct of the mediastinal lymph node was collected in premature lambs before and after delivery by cesarean section. Mean lymph flow in 7 lambs with histologically verified HMD increased progressively over 4 h after delivery to 3 times the fetal value, while lymph flow in 7 lambs without HMD increased to 3.5 times at 1 h and decreased thereafter. At 4 h after birth, lung lymph flow was significantly higher in lambs with HMD than in lambs without HMD (0.70 +/- 0.15 (SEM) vs 0.43 +/- 0.07 ml.h-1.kg-1). Lymph/plasma concentration ratio for small endogenous protein fractions (effective molecular radius, 3.6 and 3.8 nm) was significantly higher in lambs with HMD than in lambs without HMD at 2-4 h. Postmortem extravascular lung water was significantly higher in lambs with HMD (6.1 +/- 0.5 vs 4.3 +/- 0.3 ml/g dry lung weight). It is concluded that lung water is high in lambs with HMD, which appears to be a result both of delayed absorption of fetal lung liquid and increased permeability of the pulmonary exchange vessels.