Albumin Clearance Research Articles

Assessment of Human Endothelial PermeabilityIn Vitro: The Importance of Cell Seeding Density

In this study we have evaluated the importance of certain factors affecting the permeability of confluent monolayers of human umbilical vein endothelial cells (HUVEC) to albumin. Inclusion of serum in the assay medium and fibronectin-pretreatment of the membrane supports on which the monolayers were grown decreased basal permeability. Most importantly, the endothelial cell (EC) seeding density was demonstrated to be a crucial determinant of basal permeability. In this study, the lowest % clearance of albumin was obtained with a seeding density of 6.0 × 105 cells/cm2 (% clearance = 2.90 ± 0.08%). In contrast, lower (4.8 × 105 cells/cm2) or higher (9.91 × 105 cells/cm2) seeding densities resulted in significantly (P < 0.05) higher clearances. Permeability was increased on stimulation with tumour necrosis factor-α, interleukin-1α/β or interferon-γ and the fold increases were demonstrated to be dependent on the initial HUVEC seeding density. These results demonstrate the importance of assessing the effects of...

Impact of Lisinopril and Atenolol on Kidney Function in Hypertensive NIDDM Subjects With Diabetic Nephropathy

Diabetic nephropathy is characterized by hypertension and a relentless decline in kidney function. Angiotensin-converting enzyme inhibitors have been claimed to preserve kidney function better than an equal blood pressure (BP) reduction with conventional antihypertensive treatment (renoprotection). We compared the effect on kidney function of lisinopril (10-20 mg/day) and atenolol (50-100 mg/day) in hypertensive NIDDM patients (mean age 60 +/- 8 years) with diabetic nephropathy. Forty-three (21 lisinopril and 22 atenolol) patients were enrolled in a 1-year randomized double-blind parallel study. Eight patients dropped out, and the results for the remaining 35 patients (16 lisinopril and 19 atenolol) are presented. Diuretics were required in 10 of 16 lisinopril patients and 12 of 19 atenolol patients. The following variables were measured: 24-hour ambulatory BP (Takeda TM2420), albuminuria (enzyme-linked immunosorbent assay), fractional albumin clearance, and glomerular filtration rate (GFR) ([51Cr]EDTA technique). The average reduction in mean arterial BP during the 12 months was identical in the two groups 12 +/- 2 vs. 11 +/- 1 mmHg in the lisinopril and atenolol group, respectively. Albuminuria was on average reduced 45% in the lisinopril group vs. 12% in the atenolol group (P < 0.01), and fractional albumin clearance was on average reduced 49% in the lisinopril group vs. 1% in the atenolol group (P < 0.05). GFR declined identically in the two groups 11.7 +/- 2.3 vs. 11.6 +/- 2.3 ml.min-1.year-1 in the lisinopril and atenolol groups, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Antihistaminergic pretreatment prevents tissue extravasation of albumin from intra-abdominal trauma in rats.

Intra-abdominal surgery causes a loss of plasma into tissues within and around the abdomen, endangering tissue viability. Mast cells, containing histamine, are abundant in the abdominal cavity. In a model of mechanical intra-abdominal trauma, we investigated whether pretreatment with histaminergic H1 and H2 receptor blockers counteracts this extravasation. In Wistar rats under chloralose anaesthesia, tissue clearances of labelled albumin were determined by a double isotope technique. Four groups were studied: Traumatized rats, pretreated (n = 10) and non-pretreated (n = 9): non-traumatized rats; pretreated (n = 10) and non-pretreated (n = 9). In the traumatized rats, given pyrilamine 10 mg kg-1 (H1) plus cimetidine 25 mg kg-1 (H2) just before the trauma, the blockers prevented the haemoconcentration from loss of plasma and the drop in arterial pressure during the very trauma procedure, observed among non-pretreated rats. Furthermore, they significantly decreased the clearance of albumin in the abdominal wall and the pancreas. In the non-traumatized animals, the blockers lowered arterial pressure and heart rate. In conclusion, the anti-histaminergic pretreatment decreased the trauma induced leakage of albumin, by mechanisms which may involve readjustments of pressures and flows in capillaries as well as a prevention of histamine effects on capillary permeability.

Renal and microvascular effects of an aldose reductase inhibitor in experimental diabetes: Biochemical, functional and ultrastructural studies

Aldose reductase inhibitors, and particularly sorbinil, have been reported to prevent glomerular basement membrane thickening (GBMT) and albuminuria development in diabetic rats, but contradictory observations have been published. The aim of this study was to answer the following questions (i) is the corrective effect of sorbinil on GBMT, if confirmed, associated with an effect on collagen metabolism alterations? (ii) Is it associated with an effect on microvascular functional alterations? We therefore studied the influence of sorbinil on glucosyl-galactosyl-hydroxylysyl-glucohydrolase activity (GGHG; EC 3.2.1.107 which is involved in the catabolism of collagen disaccharide units), 3- and 4-hydroxyproline content and GBMT by ultrastructural morphometry in the kidney cortex of streptozotocin-diabetic rats after 5 months of disease. In parallel, the effects on albumin renal clearance and another functional alteration, the microvascular response to norepinephrine, were evaluated. We confirmed a corrective effect of sorbinil on both renal albumin clearance and GBMT. In the diabetic rats, sorbinil diminished the 3-hydroxyproline (but not the 4-hydroxyproline) content, whether expressed per mg protein or per total kidney cortex relative to body weight. Sorbinil reduced GGHG activity measured in the dialysed 10,000 g supernatant whether expressed per mg protein or per total kidney cortex; this activity has been shown to be increased in diabetes. Sorbinil also corrected the microvascular response to norepinephrine which is altered in diabetes.

Isoproterenol decreases protein permeability in edematous isolated rabbit lungs: estimation of PS and sigma

The objective of the present study was to determine whether the ability of the beta-adrenergic agonist isoproterenol to attenuate pulmonary edema occurs via a permeability and/or hemodynamic mechanism. In isolated perfused rabbit lungs, the restrictive property of the vascular barrier to the movement of fluid and protein was assessed by measurements of the capillary filtration coefficient (Kf) and the transvascular clearance of 125I-labeled albumin, respectively. Regression analysis of albumin clearance vs. transvascular fluid flux was performed to estimate the permeability-surface area product (PS) and the reflection coefficient (sigma) by use of the linear or nonlinear flux equation. Arterial, capillary, and venous pressures and resistances, weight gain, and the wet-to-dry weight ratio were also assessed. Isoproterenol (8 ng.ml-1.min-1) attenuated the arachidonic acid (4 mg)-induced increases in fluid flux, wet-to-dry weight ratio, albumin clearance, and PS and the decrease in sigma. Isoproterenol had no effect on the increase in Kf, and there was no correlation between capillary pressure and fluid flux in any of the four groups. Regression analysis revealed that the non-linear flux equation provided estimates of PS and sigma that more accurately described the statistical differences in albumin clearance among the groups studied than the linear flux equation. These findings demonstrate that isoproterenol attenuated the increased transvascular flux of albumin in edematous lungs by modifying the protein permeability of the vascular barrier.

Circulating and synovial fluid hyaluronan levels

To assess the effect of intraarticular (IA) corticosteroid on hyaluronan (HA) concentrations in synovial fluid (SF) and serum and the clearance of 131I-labeled albumin from the joints of patients with rheumatoid arthritis (RA), osteoarthritis (OA), and ankylosing spondylitis (AS). SF and serum were collected before and 2 weeks and 2 months after IA steroid injection. The HA concentration was assessed using an enzyme-linked immunosorbent assay and 131I-albumin clearance from joints was assessed using an external gamma counter. In RA patients, HA concentrations in the SF were increased following IA steroids, while the serum concentrations were decreased. In OA patients, HA concentrations in SF tended to increase initially (decreasing thereafter), and were associated with increased HA concentrations in serum. There were less marked alterations in the AS patients. Albumin clearance rates were decreased significantly (2 weeks postinjection) only in the RA patients. Estimated HA flux revealed discrepancies between the HA concentration and the rate of flux in RA and AS patients. These findings suggest that IA steroid injection is associated with a restoration in the relationship between SF and serum HA concentrations toward normal levels.

Treatment of chronic malodorous atrophic ozena with ozena-vaccine

To study the effect of atrial natriuretic peptide (ANP) on vascular permeation of albumin in the fetus, ANP (167-600 ng/min) was infused into eight ovine fetuses and saline vehicle was infused into eight twin controls (gestational age 127 +/- 3 d) over a 50-min period. Using two different radiolabeled albumin markers, we determined the tissue to blood isotope ratio (TBIR), an index of albumin permeation, and the albumin clearance. Although ANP had no hemodynamic effect, a marked increase in the hematocrit was observed in ANP-infused fetuses compared with initial values (0.37 +/- 0.04 vs 0.42 +/- 0.04, p < 0.005) but was unchanged in the twin fetuses receiving saline vehicle (0.35 +/- 0.03 versus 0.35 +/- 0.02). TBIR and albumin permeation were increased in combined tissues of ANP-infused fetuses compared with saline controls (TBIR: 1.49 +/- 0.58 versus 1.29 +/- 0.3, p < 0.001; albumin clearance: 1091 +/- 1279 versus 827 +/- 1464 nL/g/min, p < 0.01). In individual tissues, TBIR was significantly increased in skin (2.88 +/- 0.67 versus 1.55 +/- 0.35, p < 0.02), muscle (1.6 +/- 0.27 versus 1.24 +/- 0.26, p < 0.02), adrenal (1.33 +/- 0.10 versus 1.13 +/- 0.15, p < 0.02), bone (1.67 +/- 0.45 versus 1.20 +/- 0.40, p < 0.02), kidney (1.52 +/- 0.25 versus 1.24 +/- 0.26, p < 0.03), and gut (1.69 +/- 0.20 versus 1.39 +/- 0.34, p < 0.03).(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of endocytosis inhibitors on alveolar clearance of albumin, immunoglobulin G, and SP-A in rabbits

Protein in the alveolar space may be cleared by endocytosis and degradation inside alveolar epithelial cells, by transcytosis across the alveolar epithelium, or by restricted diffusion through the epithelium. The relative contributions of these three pathways to clearance of large quantities of protein from the air spaces is not known. This study investigated the effects of monensin and nocodazole, agents which inhibit endocytosis in cell culture, on alveolar epithelial protein transport in anesthetized rabbits. There was evidence that monensin and nocodazole inhibited endocytosis by the alveolar epithelium in vivo. Nocodazole increased the number of vesicles in the alveolar epithelium and capillary endothelium. Monensin increased vesicle density in the endothelium. These results suggested that the inhibitors disrupted microtubules or interrupted cellular membrane traffic in the lung. Both inhibitors decreased lung parenchymal uptake of immunoreactive human albumin from the air spaces. Monensin and nocodazole inhibited albumin uptake in cultured alveolar type II cells. Monensin increased the amount of 125I-labeled surfactant protein A associated with the lungs, compared with the quantity remaining in the air space 2 h after instillation. Although the drugs decreased alveolar epithelial protein uptake, they did not decrease alveolar clearance of 125I-labeled immunoglobulin G or 131I-labeled albumin in anesthetized rabbits. Thus monensin- and nocodazole-sensitive protein-uptake pathways do not account for most alveolar protein clearance when the distal air spaces are filled with a protein solution.

Role of H1 receptors and P-selectin in histamine-induced leukocyte rolling and adhesion in postcapillary venules.

The objective of this study was to define the nature, magnitude, and mechanisms of histamine-induced leukocyte-endothelial cell interactions in postcapillary venules of the rat mesentery using intravital microscopic techniques. Superfusion of the mesentery with histamine (10(-7)-10(-5) M) resulted in a dose-related increase in the number of rolling leukocytes, a reduction in rolling velocity, and an increased clearance of FITC-labeled rat albumin from blood to superfusate. The histamine-induced recruitment of rolling leukocytes and increased albumin clearance were prevented by histamine H1 (hydroxyzine, diphenhydramine) but not H2 (cimetidine) receptor antagonists. Because histamine induces expression of the adhesion molecule P-selectin in cultured endothelial cells, a monoclonal antibody directed against rat P-selectin and soluble sialyl-LewisX oligosaccharide (the carbohydrate ligand to P-selectin) were also tested as inhibitors. Both were effective in preventing the histamine-induced recruitment of rolling leukocytes, but neither agent attenuated the increased albumin clearance. These observations suggest that (a) histamine recruits rolling leukocytes and increases albumin leakage in postcapillary venules via H1 receptor activation, (b) histamine-induced recruitment of rolling leukocytes is mediated in part by P-selectin expressed on the endothelial cell surface, and (c) the histamine-induced vascular albumin leakage is unrelated to leukocyte-endothelial cell adhesion. Our results are consistent with the view that histamine may act as a mediator of acute inflammatory reactions.

The Effect of Serum Albumin at the Start of Continuous Ambulatory Peritoneal Dialysis Treatment on Patient Survival

To analyze the effect of serum albumin using immunoturbidimetry, demographic, biochemical, and kinetic factors on survival of continuous ambulatory peritoneal dialysis (CAPD) patients. A review of prospectively collected data in a 2-year follow-up study of peritoneal transport kinetics. University medical center. Sixty-one patients, evaluated within 3 months after the start of CAPD. Covariables used in the survival analysis were plasma urea, and creatinine, albumin, hemoglobin, mass transfer area coefficient of creatinine, peritoneal albumin clearance, 4-hour peritoneal albumin loss, net ultrafiltration, age, blood pressure, body mass index, difference between actual and ideal bodyweight, and presence or absence of systemic disease. Overall survival was 64% at 2 years. Median serum albumin was 30.9 g/L, range 18.1-43.9 g/L. Patients with a serum albumin below the median had a lower survival rate than those higher than the median (2-year survival 49% vs 79%, p = 0.01). Using the Cox model, survival was related to systemic disease (p = 0.004), age (p = 0.02), hemoglobin (p = 0.03), and serum albumin (p = 0.1). The results confirm the strength of serum albumin as predictor of survival. However, in this study serum albumin merely reflected the presence of a systemic disease, which was the most important risk factor for patient survival.

Glomerular size-and charge selectivity in Type 2 (non-insulin-dependent) diabetic patients with diabetic nephropathy

In an attempt to evaluate the mechanisms of proteinuria in diabetic kidney disease, we measured the renal clearances of albumin, total IgG, and IgG4 in 20 male Type 2 (non-insulin-dependent) diabetic patients with diabetic glomerulosclerosis (biopsy proven), in 10 male Type 2 diabetic patients without nephropathy (urinary albumin excretion rate < or = 30 mg/24 h), and in 10 healthy male subjects. The fractional clearance of albumin was increased in patients with nephropathy: 659 (42-4355) x 10(-6) (median (range)), compared to 2.6 (0.2-14.2) x 10(-6) in patients without nephropathy, and 2.3 (0.4-4.2) x 10(-6) in healthy subjects. The fractional clearance of total IgG (neutral) and of IgG4 (anionic) was 40-50 times higher in patients with nephropathy compared to the two other groups. The IgG/IgG4 selectivity index was not significantly different in the three groups, being: 1.12 (0.06-5.65), 1.16 (0.45-3.72) and 1.35 (0.65-3.34) in patients with nephropathy, patients without nephropathy, and healthy subjects, respectively. The IgG/albumin selectivity index was decreased in patients with nephropathy: 0.27 (0.01-1.26) compared to 1.29 (0.07-2.67) (p < 0.05) and 1.23 (0.76-7.84) (p < 0.001) in patients without nephropathy and healthy subjects, respectively. No significant change in IgG/albumin selectivity index was observed between patients without nephropathy and healthy subjects. The systolic blood pressure was elevated in the patients with nephropathy: 164 +/- 21 mm Hg (mean +/- SD) compared to patients without nephropathy: 145 +/- 20 mm Hg (p < 0.05) and to healthy subjects: 133 +/- 19 mm Hg (p < 0.005).(ABSTRACT TRUNCATED AT 250 WORDS)

Reduced permselectivity in isolated perfused rat kidneys following small elevations of glomerular capillary pressure.

A modified rat kidney preparation was used to explore how changes in hydrostatic pressure affect the permselective properties of the glomerular capillary bed. The maximally vasodilated kidneys of 18 rats were perfused with albumin solutions (16.7 g l-1) at different flow rates and hence arterial pressures (PA). One kidney in each rat was exposed to pressure elevations with the other kidney serving as a control perfused at constant PA of about 100 mmHg. Both the vascular resistance to flow and the glomerular filtration rate (GFR 34.6 +/- 2.9 ml min-1 100 g-1) were similar in the two kidneys at equal PA and remained constant throughout the experiment. The ratio of albumin clearance over GFR (theta) was initially around 0.4% at constant PA and gradually increased during 1.5 h to reach 0.7% at the end of the experiment. A direct increase of PA from 100 to 200 mmHg for 15 min resulted in a calculated increase of the effective glomerular filtration pressure gradient of 10-15 mmHg and in a two-fold increase of theta when measured at an identical PA of 100 mmHg. Albumin clearance was almost fully normalized within 20 min similar to that observed in e.g. skeletal muscle. However, the glomerular capillary barrier seemed to be far more sensitive to elevations of hydrostatic pressure than other capillary walls which require capillary pressure increments of 60 mmHg in order to induce similar reversible changes in permeability. Therefore, we conclude that an elevated PGC per se induces changes of glomerular permselectivity, which may have important pathophysiological implications during conditions of proteinuria.

The biologic window for chimeric l6 radioimmunotherapy

There has been little success in using radioimmunotherapy in patients with adenocarcinoma, partly because of the low tumor uptake of the administered monoclonal antibody (MoAb). The authors recently reported therapeutic response in advanced cancer patients who received 131I chimeric-L6 MoAb. The L6 MoAb identifies abundant, nonshed antigen that is expressed in many human carcinomas, including carcinomas of the lung, breast, colon, and ovary. In vitro both mouse L6 (L6) and chimeric L6 (ChL6) mediate complement-dependent tumor cytolysis with human serum, and antibody-dependent tumor cell cytolysis with normal human peripheral blood mononuclear cells. The authors have used L6 or ChL6 for radioimmunotherapy to exploit their biologic activity to create a "therapeutic window" of increased vascular permeability, allowing more 131I MoAb to reach the tumor. A reactive target is present in the vascular endothelium but can be covered by unlabeled L6 or ChL6. Nine patients with metastatic breast cancer were treated on a therapy protocol and received imaging and therapy doses of 131I ChL6 on two sequential days at 4 week intervals. During each treatment cycle, serum cytokines, complement, albumin, and 131I ChL6 blood clearance were monitored, peripheral blood mononuclear cell activation was assessed, and tumor uptake and response were documented. After L6 or ChL6 was infused, patients demonstrated immediate serum-complement activation, manifested by rapidly decreasing levels of serum complements 3 and 4. Tumor uptake of the second 131I MoAb (therapeutic) injection, given after the second daily injections of 200 mg MoAb, was usually higher than the tumor uptake of the first 131I MoAb (imaging) dose given after a single 200 mg infusion of MoAb. Although serum complement frequently decreased after the first 50-100 mg dose of L6 or ChL6, elevation of soluble interleukin-2 receptor (IL-2R) in serum was seen only in patients who received 150 mg or more of L6 or ChL6. In the nine treated patients, with only one exception, the higher grade of therapeutic tumor response was seen in patients with a greater increase in IL-2R levels. The clinical importance of understanding these mechanisms is emphasized by the occurrence of measurable tumor regressions in five of the first nine advanced metastatic breast cancer patients treated in this manner. Absence of pulmonary edema and delayed release of dose-dependent IL-2R suggest that targeting of the pulmonary endothelium by L6 or ChL6 is not the major cause of the observed biologic effects. This unique response of a solid tumor to radioimmunoconjugate therapy may be secondary to both the increased delivery of the radioimmunoconjugate to tumor cells caused by enhanced vascular permeability, and to synergistic effects of radiation and activated effector cell mechanisms.

Analysis of the pressure-flow characteristics of isolated perfused rat kidneys with inhibited tubular reabsorption.

The renal hemodynamics were studied in an isolated perfused rat kidney model modified for investigations of the glomerular permeability characteristics. The tubular reabsorptive activity was inhibited by perfusion at low temperature (8 degrees C) in the presence of furosemide and nitroprusside resulting in a dramatic increase in the filtered load of fluid and solute reaching the tubules and hence in tubular pressure. The glomerular filtration rate (GFR), arterial pressure (PA) and needle pressure (intrarenal tissue pressure, PiR) were continuously recorded and the glomerular hydrostatic pressure was estimated by an arterial occlusion technique. The pre- to postglomerular resistance ratio was calculated from the pressure vs. GFR relationships for two perfusates having differing oncotic pressures (pi = 5.5 and pi = 20 mmHg), from which estimations of glomerular hydrostatic pressures (PGC) were concomitantly made. Thus, increases in delta pi could be exactly counterbalanced by equally large increases in PGC for any given GFR, the needle and Bowman's capsule pressures being dependent on GFR but not on plasma colloid oncotic pressure. The experimental interventions resulted in a pronounced elevation of PiR as compared with in vivo conditions, while the PGC values were in a normal range, resulting in reduced glomerular filtration pressures. Furthermore, the clearance of albumin varied with the oncotic pressure in agreement with the notion of heteroporosity.

Release of ED1 fibronectin from matrix of perfused lungs after vascular injury is independent of protein synthesis.

Fibronectin (Fn) is an adhesive protein found in the plasma and extracellular tissue matrix. Locally synthesized tissue or cellular Fn (cFn) has extra domains (ED1 and ED2) not present in liver synthesized plasma Fn (pFn). In the lung, Fn is found in the endothelial and epithelial basement membranes, as well as in the interstitial matrix. Utilizing murine monoclonal antibodies to ED1 of cFn, we studied the release of total Fn as well as ED1-Fn into the plasma-free perfusate of the isolated perfused rabbit lung in relation to changes in lung weight due to fluid accumulation after oxidant (H2O2) challenge. Both parameters were also studied after addition of cycloheximide (20 micrograms/ml perfusate) to the perfusion medium to inhibit lung protein synthesis. After continuous H2O2 challenge (11 nmol.ml buffer-1.min-1), there was a 2.25 +/- 0.62 g increase in lung weight over 60 min. Measurement of 125I-labeled albumin clearance at 20 min after the start of H2O2 infusion confirmed an increase in lung endothelial protein permeability after H2O2 treatment. Fn antigen was released into the perfusate as early as 15 min after oxidant challenge. By 60 min, total perfusate Fn increased in H2O2-treated lungs (n = 6) to 2.10 +/- 0.48 micrograms/ml compared with only 0.35 +/- 0.09 micrograms/ml in normal control lungs (n = 5). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of nonreduced samples revealed that the Fn released consisted of primarily intact (440 kDa) Fn as well as Fn fragments. A rapid release of ED1-Fn paralleled the increased release of total Fn.(ABSTRACT TRUNCATED AT 250 WORDS)

Preservation of Renal Haemodynamic Response to an Oral Protein Load in Non‐insulin‐dependent Diabetes Mellitus

Glomerular Filtration Rate (GFR) and Effective Renal Plasma Flow (ERPF) were determined, for 2 h prior to and 3 h following the ingestion of a 1.2 g kg-1 meat meal, in seven normotensive normoalbuminuric Type 2 diabetic patients exhibiting good glycaemic control (fasting plasma glucose (mean +/- SD): 7.2 +/- 2.0 mmol l-1; glycosylated haemoglobin: 8.1 +/- 1.7%) and in nine normal subjects selected for similar basal GFR values. Baseline GFR and ERPF (corrected to 1.73 m2 surface area) were 83 +/- 10 and 410 +/- 76 ml min-1 for the Type 2 diabetic patients and 86 +/- 11 and 405 +/- 113 ml min-1 for the normals. GFR increased by 38 +/- 8 and 32 +/- 15% in the diabetic patients and normals, to 108 +/- 25 and 105 +/- 26 ml min-1 (p < 0.01 vs baseline). Peak ERPF was 501 +/- 127 and 476 +/- 119 ml min-1 for the two respective groups (p < 0.01 vs baseline). Filtration fractions at peak GFR and EPRF values were unchanged from baseline for either groups. Fractional clearance of albumin for the Type 2 diabetic patients was unaltered by protein ingestion. Therefore, protein ingestion in Type 2 diabetes, as in normals, results in an acute elevation of GFR. Absolute and incremental changes in GFR were identical for the two groups. These data demonstrate a preserved capacity for renal vasodilatation in Type 2 diabetic patients despite their greater chronological age.

Isoproterenol antagonizes endothelial permeability induced by thrombin and thrombin receptor peptide

We determined whether 1) amino-terminal peptides of the thrombin receptor increase endothelial permeability to a comparable extent as alpha-thrombin does, 2) isoproterenol attenuates the thrombin-induced increase in endothelial permeability by an antagonistic action to that of thrombin or by lowering baseline permeability, and 3) isoproterenol decreases permeability via stimulation of the beta 2-adrenergic receptor. Permeability across monolayers of bovine pulmonary artery endothelial cells (CCL 209) was assessed by the clearance of 125I-labeled albumin. Thrombin receptor peptides increased permeability at 1 microM but required a dose of between 10 and 100 microM to equal the permeability response of 1 microM alpha-thrombin. Dose-response experiments demonstrated that isoproterenol antagonized the action of alpha-thrombin and a thrombin receptor peptide on endothelial permeability and that it lowered baseline permeability. This permeability-decreasing action of isoproterenol occurred via stimulation of the beta 2-adrenergic receptor. Terbutaline, a partial beta 2-agonist, prevented the thrombin-induced permeability, but dobutamine, a partial beta 1-agonist, did not. The active stereoisomer of terbutaline and the racemic form mimicked the action of isoproterenol, but the inactive stereoisomer had no effect. ICI-118,551, a specific beta 2-receptor antagonist, prevented the permeability-decreasing action of isoproterenol, whereas ICI-89,406, a specific beta 1-receptor antagonist, did not. Competitive binding studies of 125I-pindolol with ICI-118,551 or ICI-89,406 demonstrated the presence of beta-adrenergic receptors, predominantly beta 2-receptors, on cell membrane homogenates.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of angiotensin-converting enzyme reduces urinary albumin excretion but not regional albumin clearance in experimental diabetes

Albuminuria and increased regional albumin clearance are known to develop concomitantly in diabetes. The anti-proteinuric effect of angiotensin-converting enzyme inhibitor therapy is well established. We studied whether angiotensin-converting enzyme inhibitor therapy with perindoprilat in comparison with conventional antihypertensive treatment could influence the development of increased regional albumin clearance as well as albuminuria in experimental diabetes. Rats with streptozotocin-induced diabetes were randomized into a saline group (n = 7), a perindoprilat (1 mg/kg per day) group (n = 8), and a hydralazine (3 mg/kg per day) group (n = 6); six rats served as non-diabetic controls. After 6–8 weeks, blood pressure was equally reduced in the perindoprilat- and hydralazine-treated groups (P < 0.01). Twenty-four-hour urinary protein and albumin excretion were increased in diabetic rats compared to control rats (P < 0.001). Hydralzine did not reduce 24 h protein or albumin excretion, whereas perindoprilat treatment reduced both (P < 0.001) to levels comparable to those of control rats. Regional albumin clearance, assessed in the eye, ileum, lung, skeletal muscle and skin, was clearly elevated in diabetic rats compared to control rats; however, neither drug therapy had an effect on albumin clearance.

Incorporation of fibronectin into matrix decreases TNF-induced increase in endothelial monolayer permeability

Plasma fibronectin, a dimeric adhesive protein in blood, incorporates into the subendothelial and interstitial matrix in the lung especially during vascular injury. Fibronectin in the matrix is believed to influence cell-cell interaction and endothelial cell adhesion to the collagen-rich extracellular matrix. We previously observed that addition of purified soluble human plasma fibronectin (hFn) to cultured pulmonary endothelial monolayers attenuates the increase in protein permeability of such monolayers exposed to tumor necrosis factor-alpha (TNF-alpha). In the current study, we determined the specificity of this permeability response to fibronectin by comparing hFn to two other purified adhesive proteins in human plasma, i.e., vitronectin (Vn) and fibrinogen (Fg). We also determined whether matrix incorporation was essential for this hFn-mediated protective response by comparing normal intact hFn to either hFn alkylated with N-ethylmaleimide (NEM) or to purified 160/180-kDa hFn fragments, since these alternate forms of fibronectin are believed to exhibit limited ability to incorporate into matrix. Calf pulmonary artery endothelial (CPAE) monolayers (3-4 days postseeding) were exposed to human recombinant TNF-alpha for 18 h at a medium concentration of 200 U/ml followed by assessment of protein permeability using transendothelial 125I-labeled albumin clearance. Dimeric hFn (600 micrograms/ml) significantly (P < 0.05) reduced the TNF-induced increase in endothelial monolayer permeability. Vn or Fg, added at equal molar concentrations to the hFn, were unable to attenuate endothelial permeability. Immunofluorescent analysis utilizing antibodies specific to either hFn, human Vn, or human Fg revealed incorporation of the exogenous hFn into the extracellular matrix, but no matrix incorporation of Vn or Fg. Both NEM-treated dimeric hFn as well as purified 160/180-kDa fragments of hFn, which cannot incorporate into the matrix, were also unable to prevent the TNF-induced increase in protein permeability. Thus the ability for soluble hFn to reduce the TNF-induced increase in lung endothelial monolayer permeability was specific and dependent on its incorporation into the extracellular matrix.

Continuous Ambulatory Peritoneal Dialysis with a High Flux Membrane

The standard peritoneal equilibration test (PET) was performed in 66 patients on continuous ambulatory peritoneal dialysis (CAPD). Patients were classified as low (n = 5), low average (n = 22), high average (n = 27), and high (n = 12) transporters based on the dialysate/plasma creatinine (D/P Cr) after 4 hr dwells. After an average time interval of 14 months on CAPD, indices of dialysis adequacy and nutrition were assessed. Based on monitoring of patient chemistries and drain volumes, peritoneal transport was considered stable during the interval. Instilled volumes and exchange tonicity were individualized in each patient to achieve combined renal and dialysis weekly creatinine clearance and KT/V urea that were not significantly different between groups. High transporters had significantly (p < 0.05) lower mean serum albumin, net protein catabolic rate (nPCR), lean body mass calculated from creatinine kinetics, and daily creatinine production (and presumably lower muscle mass), and higher albumin clearances compared to one or more lower transport groups. In conclusion, we hypothesize that high transporters are prone to protein malnutrition related to increased dialysate protein losses, and perhaps suppression of appetite, with increased use of hypertonic exchanges. High transporters are candidates for protein supplementation on CAPD or transfer to nightly intermittent peritoneal dialysis where short cycles provide more ultrafiltration with less glucose absorption.