Solvent Drag Reflection Coefficient Research Articles

VEGF increases paracellular transport without altering the solvent-drag reflection coefficient

Vascular endothelial growth factor (VEGF) increases microvascular permeability and has been implicated in the development of numerous pathologies including diabetic retinopathy (DR), hypoxia/ischemia, and tumor biology. The transport pathways by which water and solutes cross the endothelium in response to VEGF, however, are not completely understood. We measured, in real time, bovine retinal endothelial cell (BREC) hydraulic conductivity (Lp), 70 kDa dextran permeability (Pe), and the solvent-drag reflection coefficient ( σ) before and after addition of 50 ng/ml VEGF. The diffusional permeability coefficient for dextran (Pd) was measured before pressure gradient application. The sudden application of a 10-cm H 2O hydrostatic pressure gradient induced water and solute fluxes that decayed to steady-state values after approximately 2 h. Subsequently, the addition of VEGF significantly increased Lp and Pe by 4.3-fold ± 0.7-fold and 3.0-fold ± 0.3-fold, respectively, after 110 min; however, the reflection coefficient remained approximately constant throughout the experiment (approximately 0.8). These observations suggest that water and dextran utilize common paracellular channels across BREC monolayers. Furthermore, the addition of VEGF increases the number or availability of channels but does not alter the selectivity of the monolayer to 70 kDa dextran.

Effect of venous air embolization on pulmonary microvascular protein permeability.

An increase in pulmonary lymph flow and lymph protein clearance following pulmonary air embolization has been interpreted as evidence of increased pulmonary microvascular permeability. The authors hypothesized that air embolization does not alter the pulmonary microvascular permeability to protein and that this could be demonstrated by determining the effect of air embolization on the pulmonary solvent drag reflection coefficient (sigma(f)). Anesthetized dogs were instrumented with left atrial balloon-tipped catheters, pulmonary lymphatic cannulae, and inferior vena caval catheters. Values were determined for pulmonary lymph flow (Q(L)) and the lymph-to-plasma protein concentration ratio (C(L)/C(P)) at baseline, after C(L)/C(P) was decreased to a filtration independent value by raising left atrial pressure via progressive balloon inflation and after 2 h of air embolization into the inferior vena cava with continued left atrial hypertension. Q(L) increased and C(L)/C(P) decreased to a filtration-independent value following induction of left atrial hypertension. Air embolization induced during left atrial hypertension resulted in no significant change in C(L)/C(P). The authors were unable to demonstrate that sigma(f) changed following pulmonary air embolization. Utilizing the washdown technique, the authors could not find any evidence that pulmonary microvascular protein permeability is altered by air embolization.

A transmural pressure gradient induces mechanical and biological adaptive responses in endothelial cells.

A sudden increase in the transmural pressure gradient across endothelial monolayers reduces hydraulic conductivity (L(p)), a phenomenon known as the sealing effect. To further characterize this endothelial adaptive response, we measured bovine aortic endothelial cell (BAEC) permeability to albumin and 70-kDa dextran, L(p), and the solvent-drag reflection coefficients (sigma) during the sealing process. The diffusional permeability coefficients for albumin (1.33 +/- 0.18 x 10(-6) cm/s) and dextran (0.60 +/- 0.16 x 10(-6) cm/s) were measured before pressure application. The effective permeabilities (measured when solvent drag contributes to solute transport) of albumin and dextran (P(ealb) and P(edex)) were measured after the application of a 10 cmH(2)O pressure gradient; during the first 2 h of pressure application, P(ealb), P(edex), and L(p) were significantly reduced by 2.0 +/- 0.3-, 2.1 +/- 0.3-, and 3.7 +/- 0.3-fold, respectively. Immunostaining of the tight junction (TJ) protein zonula occludens-1 (ZO-1) was significantly increased at cell-cell contacts after the application of transmural pressure. Cytochalasin D treatment significantly elevated transport but did not inhibit the adaptive response, whereas colchicine treatment had no effect on diffusive permeability but inhibited the adaptive response. Neither cytoskeletal inhibitor altered sigma despite significantly elevating both L(p) and effective permeability. Our data suggest that BAECs actively adapt to elevated transmural pressure by mobilizing ZO-1 to intercellular junctions via microtubules. A mechanical (passive) component of the sealing effect appears to reduce the size of a small pore system that allows the transport of water but not dextran or albumin. Furthermore, the structures of the TJ determine transport rates but do not define the selectivity of the monolayer to solutes (sigma).

EFFECT OF ONO-5046, A SPECIFIC NEUTROPHIL ELASTASE INHIBITOR, ON THE PHORBOL MYRISTATE ACETATE-INDUCED INJURY IN ISOLATED DOG LUNG

Phorbol myristate acetate (PMA) activates neutrophils and causes acute lung injury. We determined the effect of ONO-5046, a specific neutrophil elastase inhibitor, on the increase in microvascular permeability induced by PMA in isolated dog lung perfused with autologous blood at a constant perfusion flow. The vascular permeability was assessed by the capillary filtration coefficient (Kf,c) and the solvent-drag reflection coefficient (sigmaf). PMA (13.3mug) increased vascular permeability, as evidenced by an increase in Kf,c from 0.18 +/- 0.02 to 0.92 +/- 0.14mL/min/cmH2O/100g and a decrease in sigmaf to 0.35 +/- 0.01 as compared to control values of 0.69 +/- 0.06. The PMA-induced changes in Kf,c and sigmaf were dose-dependently attenuated by pretreatment with ONO-5046 (2-20mg). We conclude that ONO-5046 can effectively attenuate the PMA-induced injury in the isolated blood-perfused dog lungs.

Skeletal muscle ischemia-reperfusion causes transitory increase in microvascular protein permeability.

The aim of this study was to determine whether the length of ischemia in skeletal muscle influences the return of normal microvascular permeability during reperfusion in addition to influencing the size of the initial changes. In anesthetized rabbits, the transvascular clearance of labeled albumin was measured in the gastrocnemius and soleus muscles during the first, second, third, or fourth hour of reperfusion after 1, 2, 3, or 4 h of ischemia. The size of the increases in albumin clearance, tissue water, and myeloperoxidase activity during the first hour of reperfusion was dependent on the length of ischemia. The return of the albumin clearance to control values during the fourth hour of reperfusion was independent of the length of ischemia. Tissue water, extravascular mass of native albumin, and myeloperoxidase activity remained elevated during the 4 h of reperfusion. After 4 h of ischemia, the solvent-drag reflection coefficient for albumin was significantly less than control during the first hour of reperfusion. The value during the fourth hour of reperfusion was not significantly different from control. These results suggest that the inflammatory mediators producing a change in permeability are washed out of the microvasculature during the first few hours of reperfusion.

Protease inhibition attenuates microvascular dysfunction in postischemic skeletal muscle.

Neutrophils accumulate in skeletal muscle subjected to ischemia-reperfusion and appear to contribute to reperfusion-induced microvascular dysfunction. The overall objective of this study was to assess the role of the neutrophilic hydrolytic enzyme elastase in ischemia-reperfusion-induced granulocyte accumulation and microvascular dysfunction in skeletal muscle. We examined the effect of three structurally unrelated elastase inhibitors [eglin C, MeOsuc-Ala-Ala-Val-CH2Cl (MAAPV), or L-658758], administered at the onset of reperfusion, on neutrophil content and the increase in microvascular permeability induced by 4 h of ischemia and 0.5 h of reperfusion in the isolated canine gracilis muscle. Changes in vascular permeability (1 - sigma) were assessed by determining the solvent drag reflection coefficient for total plasma proteins (sigma) in the following groups: 1) 4.5 h of continuous perfusion (nonischemic), 2) ischemia-reperfusion alone, 3) ischemia-reperfusion + eglin C, 4) ischemia-reperfusion + MAAPV, and 5) ischemia-reperfusion + L-658758. Muscle neutrophil content was monitored by assessing tissue myeloperoxidase (MPO) activity in biopsies obtained at the end of the experiments. In nonischemic muscles, 1 - sigma and MPO activity averaged 0.13 +/- 0.03 and 0.7 +/- 0.2 units/g wet wt, respectively. Ischemia-reperfusion was associated with marked increases in microvascular permeability (1 - sigma = 0.39 +/- 0.02) and muscle MPO activity (8.9 +/- 1.2 units/g wet wt) that were attenuated by eglin C, MAAPV, and L-658758 (1 - sigma = 0.21 +/- 0.01, 0.22 +/- 0.02, and 0.21 +/- 0.03, respectively; MPO activity = 2.7 +/- 0.4, 2.1 +/- 0.8, and 2.8 +/- 1.8 units/g wet wt, respectively). These results suggest that granulocyte accumulation in postischemic skeletal muscle is dependent on the release of elastase from activated phagocytic cells. Moreover, neutrophilic elastase appears to play a major role in reperfusion-induced increases in microvascular permeability in skeletal muscle.

Fructose-1,6-diphosphate or adenosine attenuate leukocyte adherence in postischemic skeletal muscle.

The purpose of this study was to determine whether fructose-1,6-diphosphate (FDP) or adenosine (Ado), administered at the onset of reperfusion, would prevent ischemia/reperfusion (I-R)-induced leukocyte adherence and microvascular dysfunction in skeletal muscle. Changes in vascular permeability and tissue neutrophil content were assessed by measurement of the solvent drag reflection coefficient (delta) for total plasma proteins and muscle myeloperoxidase (MPO) activity, respectively, in continuously perfused, isolated canine gracilis muscles and in muscles subjected to I-R alone, I-R + FDP, and I-R + Ado. To determine whether FDP or Ado would attenuate leukocyte-endothelial cell adhesive interactions induced by I-R, leukocyte adherence and emigration were assessed in postischemic mouse cremaster muscles, using intravital microscopy in the presence and absence of FDP or Ado during reperfusion. I-R was associated with a marked increase in microvascular permeability and muscle MPO activity relative to nonischemic controls. These increases were attenuated by FDP and Ado. I-R also increased the number of adherent and emigrated leukocytes relative to control. I-R-induced leukocyte adherence and emigration were significantly attenuated by either FDP or Ado. These results indicate that FDP and Ado attenuate postischemic microvascular barrier dysfunction in skeletal muscle by a mechanism that may be related to their ability to inhibit leukocyte adhesion and emigration.

Solvent drag of LDL across mammalian endothelial barriers with increased permeability.

We investigated the mechanisms of hamster low-density lipoprotein (LDL) transport across the endothelial barrier in individually perfused venular microvessels in hamster mesentery. These experiments are the first to use microperfusion techniques and quantitative fluorescence microscopy to investigate LDL transport across mammalian microvessel endothelium. The apparent permeability coefficient for hamster LDL, PsLDL, rose from 2.7 x 10(-7) cm/s at control to 23.2 x 10(-7) cm/s at the peak of the biphasic increase in microvessel permeability after exposure of the vessels to 100 microM histamine. Close to the peak, PsLDL rose 1.85 x 10(-7) cm/s for every centimeter of H2O increase in hydrostatic pressure. Thus, at a mean pressure of 11.3 cmH2O, 90% of the LDL flux was coupled to transendothelial water flow by a solvent drag mechanism. The corresponding solvent drag reflection coefficient for hamster LDL was estimated to be approximately 0.8. These results are consistent with sieving hamster LDL (effective radius 14.9 nm) through equivalent pores of approximately 22 nm radius. Similar results were found with human LDL (effective radius 13.2 nm) in hamster microvessels. The results provide a bridge between studies of LDL transport across cultured endothelial barriers, where high diffusive permeability coefficients to LDL may obscure the contributions of solvent drag, and studies in whole animals, where the consequences of sieving of LDL at the vessel wall, even in the high permeability state, have not received much attention.

Identification of microvascular transport pathways in skeletal muscle

The solvent-drag reflection coefficient (sigma f) was measured from plasma disappearance (integral-mass balance method) for native albumin and four fluorescent solutes of radii from 2 to 16 nm in the isolated, plasma-perfused cat hindlimb preparation. The data for the smallest solutes were measured > 2 h after tracer addition and at high filtration rates to avoid underestimation of sigma f due to tracer diffusion. A two-pore model was fit (small-pore and large-pore radii, approximately 3.5 and 23 nm, respectively, 84% of hydraulic capacity in small pores) to these data using an objective computer-based estimation procedure. In the model, membrane sigma f was determined by flow weighting the sigma f values for the two pathways. Also, the phenomenon of volume circulation among the pathways was included. In different limbs, the permeability-surface area (PS) product was measured for the smallest solute, alpha-lactalbumin, from its perfusate-disappearance transient and a linear diffusion model. The PS value estimated was 0.11 +/- 0.026 (95% confidence limits) ml.min-1 times 100 g muscle-1. These PS values were found to be coincident with those predicted using parameter sets derived from the multiparameter 95% confidence space consistent with the two-pore model fits. The two-pore model also closely predicted PS data for small solutes from other studies in skeletal muscle; however, it failed to adequately describe small-molecule transport data from osmotic transient studies. It was necessary to add a water-exclusive pathway (40% of total hydraulic capacity) to account for these latter data; however, the predictions with this addition were still consistent with the data measured in the present study. We conclude that pore models can describe both macromolecular and small solute reflection coefficient and PS data in skeletal muscle.

Effect of high blood flow on pulmonary vascular permeability to protein.

The elevated cardiac output associated with exercise increases lung lymph flow and may increase extravascular lung water. However, it is not known if extremely elevated cardiac output alters pulmonary vascular permeability. The hematocrit-protein method was used to determine the solvent drag reflection coefficient, an index of vascular permeability to proteins, in the isolated blood-perfused canine lung lobe. Microvascular pressure was obtained by double vascular occlusion. Lobes filtered fluid during perfusion at normal flow, 0.451 +/- 0.005 l/min (LF; n = 8), or high flow, 2.319 +/- 0.080 l/min (HF; n = 7). In the LF, venous pressure was elevated to 19.0 +/- 0.5 Torr to induce filtration, whereas Pv was 3.3 +/- 0.1 Torr in the HF. In HF vs. LF, respectively, arterial pressure was 61.4 +/- 7.1 vs. 28.0 +/- 1.0 Torr (P < 0.05), microvascular pressure was 31.9 +/- 3.0 vs. 22.2 +/- 0.9 Torr (P < 0.05), and sigma was 0.52 +/- 0.07 vs. 0.51 +/- 0.02 (P > 0.05). The fivefold increase in blood flow did not alter pulmonary vascular permeability to proteins; however, the capillary filtration coefficient was fivefold greater in the HF vs. LF group (0.328 +/- 0.059 vs. 0.067 +/- 0.007; P < 0.002). These data are compatible with enzyme activity measures indicating a direct linear relationship between blood flow rate and perfused pulmonary microvascular surface area. Although the data do not rule out the possibility of increased pulmonary vascular permeability to water during very elevated blood flow rates, the greater filtration rate during elevated flow is more likely related to increases in both microvascular pressure and surface area.

Permeability of parietal pleura to liquid and proteins.

The permselectivity of the parietal pleura was determined in spontaneously breathing anesthetized rabbits and dogs. In rabbits, we injected intrapleurally 5 ml of 1-g/dl albumin solution containing 100 microCi of 131I-labeled albumin plus 100 microCi of either lactate dehydrogenase (LDH) or alpha 2-125I-macroglobulin. Dogs received 100 ml of 1-g/dl albumin solution containing 100 microCi of 131I-albumin plus 100 microCi of alpha 2-125I-macroglobulin. A transpleural pressure gradient was set, lowering the intracapsular pressure to -30 cmH2O. The solvent drag reflection coefficients (sigma f) were calculated as the ratio between tracer concentrations in capsular and pleural liquid collected at 60-180 min. In rabbits sigma f was 0.44 +/- 0.2 (SD) for albumin, 0.84 +/- 0.1 for LDH, and 0.93 +/- 0.05 for alpha 2-macroglobulin. In dogs sigma f was 0.30 +/- 0.19 for albumin and 0.53 +/- 0.15 for alpha 2-macroglobulin. The hydraulic conductivity of the parietal pleura was 2.18 +/- 1.54 microliters.h-1.cmH2O-1.cm-2 in rabbits and 1.22 +/- 1.13 microliters.h-1.cmH2O-1.cm-2 in dogs. The parietal pleura could be modeled by two pore populations with radii of 83-89 and 156-222 A. The permeability coefficient averaged 0.08-0.21 x 10(-6) cm/s for albumin, 0.06-0.09 x 10(-6) cm/s for LDH, and 0.01-0.03 x 10(-6) cm/s for alpha 2-macroglobulin.

Effect of Cold on Ischemia-Reperfusion-Induced Microvascular Permeability Increase in Cat Skeletal Muscle

The effects of reduced temperature during ischemia (I) upon microvascular permeability and resistance increases during reperfusion (R) were assessed in skeletal muscle. Protein solvent-drag reflection coefficients (σ f) and changes in vascular resistance were measured during 37°C reperfusion of isolated, whole-blood perfused cat bindlimbs after the limbs had been subjected to 3.5 h of ischemia at several temperatures. σ f was determined from the disappearance rates of water and protein from the circulating perfusate during a period of induced microvascular fluid filtration. The I/R procedure at 37°C caused σf to fall from ∼0.85 to ∼0.5, indicating a large increase in microvascular permeability. Hypothermic ischemia at 30, 22, 17, or 12°C totally abolished this drop in σ f. However, when the ischemia was at ∼5°C, there was a significant fall in σ f to ∼0.7, which was similar to the value we found previously with 5 h of continuous perfusion at this low temperature. The normothermic I/R procedure led to an increase in vascular resistance of ∼250% above the value measured prior to I/R. Hypothermic ischemia totally abolished this resistance increase, except for the lowest temperature, for which the increase was 150%. Therefore, hypothermia can prevent the microvascular dysfunction caused by 3.5 b of ischemia at 37°C in this preparation. However, when the temperature was reduced too far (∼5°C), a cold injury to the microvascular resulted in permeability and resistance increases.

Peritoneal Solvent Drag Reflection Coefficients Are within the Physiological Range

Previous estimates of the peritoneal solvent drag reflection coefficient (sigma) are widely disparate; some are outside the range expected for a semipermeable membrane (i.e., between 0 and 1). We have evaluated a novel method for determining sigma in a rabbit model of peritoneal dialysis. Test solute was infused intravenously, and sequential 2-hour isotonic and hypertonic exchanges (40 ml/kg) were performed in random order. Test solute was also added to the instilled hypertonic dialysis solution to inhibit transperitoneal solute diffusion during this exchange. Eight experiments were performed with creatinine as test solute and glucose as osmotic solute, and six experiments were performed with glucose as test solute and mannitol as osmotic solute. When using isotonic dialysis solution, the dialysate/plasma concentration ratio (D/P) for both test solutes increased throughout the exchange (p < 0.001). When using hypertonic dialysis solution, D/P creatinine was initially near 1 and decreased during the 1st hour of the exchange (p < 0.05); D/P glucose (as test solute) was initially 0.82 +/- (SEM) 0.07 and did not change during the exchange. The peritoneal diffusive permeability-area product (PA) was determined by fitting a mathematical model to the time dependence of the dialysate test solute concentration during the isotonic exchange using PA as an adjustable parameter, and sigma was determined in like manner during the hypertonic exchange using sigma as an adjustable parameter (and assuming a PA value equal to that during the isotonic exchange). The creatinine PA (1.37 +/- 0.28 ml/min) was higher than that for glucose (0.62 +/- 0.07 ml/min) as expected based on their solution diffusion coefficients. Creatinine and glucose sigma values were 0.38 +/- 0.06 and 0.43 +/- 0.05, respectively. We conclude that peritoneal sigma values for creatinine and glucose are within the physiological range.

Segmental vascular pressures in lung embolism.

Average microvascular filtration pressure and vascular permeability measures were obtained in 100-microns glass bead-embolized dog lung lobes randomly assigned to groups in which isolated perfusion was designed to produce weight gain (edema groups) or no weight gain (isogravimetric groups). The solvent drag reflection coefficient (sigma), an index of vascular permeability, was obtained during edema formation, whereas isogravimetric capillary pressure was obtained during isogravimetry. Vascular permeability increased in response to embolism, because sigma was 0.53 +/- 0.03 vs. 0.80 +/- 0.05 (P < 0.005) in embolized and control lobes, respectively. Vascular occlusion methods indicated the greatest resistance increase in response to embolism in the vascular segment represented by Pao--Pdo (arterial occlusion pressure--double occlusion pressure). Because papaverine vasodilation reduced total vascular resistance (RT; P < 0.05) by decreasing Pao (P < 0.01) without altering Pdo, the RT increase in response to embolism was likely due to both vasoconstriction and obstruction. Because Pdo approximated capillary pressure at isogravimetry, Pdo appears to estimate average filtration pressure in both embolized (n = 6) and control lungs (n = 6). Arterial pressure was 56.2 +/- 13.6 vs. 17.6 +/- 1.5 cmH2O (P < 0.01) in embolized (n = 5) and control lobes (n = 6), respectively, whereas Pdo values of 16.1 +/- 1.5 vs. 12.4 +/- 0.8 (P < 0.05) suggested relatively little increase in filtration pressure in response to embolism. If the beads obstructed 100-microns vessels, the vascular segment represented by Pao--Pdo, the major site of vasoconstriction as well as mechanical obstruction, likely includes 100-microns arteries.

Transvascular albumin flux in rabbit hindlimb after tourniquet ischemia.

Transvascular clearance of labeled rabbit albumin was measured in gastrocnemius muscle and heel skin from anesthetized rabbits after 1 or 2 h of tourniquet ischemia. Albumin clearance, calculated as a 1-h extravascular uptake, was 4.7 +/- 0.4 and 29 +/- 1 microliter.h-1 x g dry wt-1 for control gastrocnemius muscle and heel skin, respectively. During the first hour of reperfusion, the clearance was 101 +/- 23 and 56 +/- 6 microliters.h-1 x g dry wt-1 in the muscle and skin, respectively. The clearance in skin during the second and third hour of reperfusion was not different from control. The increased clearance in muscle persisted during 2 h of reperfusion but returned to control levels during the third hour of reperfusion. The solvent drag reflection coefficient was measured by increasing venous pressure during reperfusion in one leg after bilateral ischemia. In skin, the reflection coefficient, calculated as 1 minus the change in clearance divided by the change in water weight, was 0.94 +/- 0.05 and did not change with tourniquet ischemia. The reflection coefficient in muscle was 0.98 +/- 0.01 for control animals and decreased to 0.66 +/- 0.11 during the first hour of reperfusion. The reflection coefficient was not different from control during the third hour of reperfusion. The transitory increase in microvascular permeability to albumin in skeletal muscle is more indicative of an acute inflammatory response than of endothelial injury.

Effects of cold on vascular permeability and edema formation in the isolated cat limb

We investigated the effects of cold temperatures on microvascular protein permeability in the isolated constant-flow perfused cat hindlimb. The perfusates were 20% cat plasma-80% albumin-electrolyte solution (low-viscosity perfusate, approximately 1 cP) or whole blood (high-viscosity perfusate, approximately 4 cP). The time at low temperature (less than 10 degrees C) was less than 3 h (short term) or greater than 5 h (long term). Decreases in the solvent drag reflection coefficient (sigma f) indicated increases in permeability. The sigma f's were determined with the integral-mass balance method from measurement of changes in protein concentration and hematocrit induced by fluid filtration into the tissues. Short-term cold exposure did not increase permeability with either a low- or a high-viscosity perfusate, whereas long-term exposure with limb temperatures of approximately 5 degrees C significantly increased permeability when the perfusate was whole blood. In addition, we verified our previous prediction that flow had to be reduced to 6-8 ml.min-1.100 g-1 to avoid the hydrostatic edema caused by short-term perfusion with whole blood at approximately 5 degrees C. Also, we found that at approximately 3 degrees C histamine's permeability-increasing effect was totally abolished, whereas at approximately 20 degrees C this effect was partially inhibited. Hence, constant-flow perfusion at low temperature with whole blood can cause edema by a pressure-dependent mechanism, whereas long-term perfusion with this perfusate at low temperatures can cause a permeability increase that further compounds edema formation. Histamine is not responsible for this permeability increase.

Effect of extreme elevations in venous pressure on reflection coefficient in the lung

We determined whether the solvent drag reflection coefficient ( σ f) for total proteins of a canine perfused left lower lung lobe (LLL) preparation decreases at elevated venous pressures ( P v). We found that σ f (estimated using the hematocrit-protein technique) remained constant at all P v's (30–95 mm Hg) evaluated. These results were unanticipated, since previous studies reported increases in protein permeability at P v's within this range. We conducted two additional studies to better understand the basis for these observations. In the first, we evaluated the effect of high P v (85 mm Hg) on σ f of a canine perfused forelimb preparation and found σ f to be reduced. This difference in response suggests that the normal σ f's observed in the LLL were not due to high P v per se, but rather that there is some intrinsic difference between the pulmonary and the systemic circulations that accounts for the difference. The second study was designed to determine whether the normal σ f's observed in the LLL at high P v's provide meaningful information about pulmonary vascular endothelial permeability. We damaged LLL's with alloxan, oleic acid, or HCl and obtained near normal estimates of σ f at high P v. These results indicated that it is not possible to easily distinguish between a normal and a damaged pulmonary vasculature when σ f is measured at high P v. We suggest that the normal estimates of σ f obtained at high P v in the LLL result from an increased fraction of the transvascular flow occurring through pathways that exclude macromolecules.

Pressure-dependent increase in lung vascular permeability to water but not protein.

Simultaneous measures of vascular permeability to fluid (capillary filtration coefficient, Kf) and to plasma proteins (solvent drag reflection coefficient, sigma) were obtained over venous pressures (Pv) from 14 to 105 Torr in the isolated ventilated canine lung lobe (n = 70) pump perfused with autologous blood. The sigma was obtained from the relative increase in the concentration of plasma proteins vs. erythrocytes during fluid filtration. Kf's were obtained from two gravimetric methods as well as from change in hematocrit. All Kf's increased (P less than 0.05) as Pv was increased. However, sigma averaged 0.59 +/- 0.01 (range 0.54-0.67) and was unchanged (P greater than 0.05) by elevation of Pv over 20-105 Torr. In 44 lobes where all three Kf measures were obtained, gravimetric measures of Kf did not differ (P greater than 0.05) and were highly correlated with Kf obtained from hematocrit change, Vf Kf (P less than 0.001). However, both weight-based Kf's exceeded Vf Kf (P less than 0.05), suggesting that fluid filtration was overestimated by rate of lung weight gain or underestimated by hematocrit change. Increased permeability to water but not to protein over Pv from 20 to 105 Torr indicates that permeability to both can change independently and is counter to the theory that elevated vascular pressure "stretches" vascular pores.

Transport parameter estimation from lymph measurements and the Patlak equation.

Two methods of estimating protein transport parameters for plasma-to-lymph transport data are presented. Both use IBM-compatible computers to obtain least-squares parameters for the solvent drag reflection coefficient and the permeability-surface area product using the Patlak equation. A matrix search approach is described, and the speed and convenience of this are compared with a commercially available gradient method. The results from both of these methods were different from those of a method reported by Reed, Townsley, and Taylor [Am. J. Physiol. 257 (Heart Circ. Physiol. 26): H1037-H1041, 1989]. It is shown that the Reed et al. method contains a systematic error. It is also shown that diffusion always plays an important role for transmembrane transport at the exit end of a membrane channel under all conditions of lymph flow rate and that the statement that diffusion becomes zero at high lymph flow rate depends on a mathematical definition of diffusion.

Phalloidin attenuates postischemic neutrophil infiltration and increased microvascular permeability.

The aim of this study was to determine whether phalloidin (1 microM) or antamanide (1 microM), cyclic peptides that stabilize dense peripheral band and stress fiber F-actin in endothelium, would attenuate the increase in microvascular permeability induced by 4 h of ischemia and 30 min of reperfusion (I/R) in the isolated canine gracilis muscle. Changes in microvascular permeability (1 - sigma) were assessed by determining the solvent drag reflection coefficient for total plasma proteins (sigma) in muscles subjected to 4.5 h of continuous perfusion (nonischemic controls), I/R alone, I/R + phalloidin, or I/R + antamanide. Muscle neutrophil content was assessed by determination of myeloperoxidase (MPO) activity in tissue samples obtained at the end of the experiments. Fluorescent detection of nitrobenzoxadiazole-phallicidin in endothelial cell monolayers confirmed that phalloidin enters these cells. I/R was associated with marked increases in microvascular permeability and muscle neutrophil content (1 - sigma = 0.45 +/- 0.07; MPO = 8.9 +/- 0.5 units/g) relative to control (4.5 h continuous perfusion) preparations (1 - sigma = 0.12 +/- 0.03; MPO = 0.5 +/- 0.8 unit/g). These I/R-induced changes were largely prevented by administration of phalloidin (1 - sigma = 0.19 +/- 0.02; MPO = 0.8 +/- 0.4 U/g) or antamanide (1 - sigma = 0.07 +/- 0.11; MPO = 0.9 +/- 0.3 unit/g) at reperfusion. Similar results were obtained when phalloidin was administered before ischemia (1 - sigma = 0.24 +/- 0.04; MPO = 1.2 +/- 1.0 units/g). Although antamanide decreased superoxide production (by approximately 60%) and adherence to plastic (by approximately 75%) by activated neutrophils in vitro, phalloidin failed to alter these aspects of granulocyte function.(ABSTRACT TRUNCATED AT 250 WORDS)