Effects of colloid or crystalloid solutions on edemagenesis in normal and thrombomicroembolized lungs.

We studied the effects of crystalloid (75 ml/kg of Ringer's lactate) or colloid (6% dextran-70, 6% hydroxyethyl starch, or 25 ml/kg of 5% human serum albumin) fluid infusions or no treatment (control) on the calculated lung capillary (Pc)-plasma oncotic pressure (pi c) gradient and pulmonary edema. Two sets of mongrel dogs were studied: uninjured (n = 25; 14 to 22 kg) and pulmonary fibrin-microembolized (n = 25; 15 to 23 kg). In both sets of experiments, left atrial pressure was controlled (15 mm Hg) throughout the 4-h plus experimental period. In the uninjured set, the Pc-pi c gradient averaged +1.0 and -0.2 mm Hg in the hydroxyethyl starch and dextran groups, +0.7 and +2.3 mm Hg in the human serum albumin and control groups, and +6.2 mm Hg in the Ringer's lactate group. In the fibrin-microembolized group, this gradient averaged 2.6, 2.4, 3.0, 5.3, and 9.5, respectively. The extravascular lung water to bloodless dry lung wet weight ratios in the no-fluid treatment group of the uninjured and microembolism groups with increased pressure (3.8 +/- 0.3 to 4.1 +/- 0.4 [SEM]; NS) are consistent with interstitial or perivascular edema. There were, however, no significant differences noted between the respective control groups or any fluid treatment group in either set of experiments. These data support the view that infusion of either colloid or crystalloid solutions in normal or pulmonary fibrin-microembolized lungs does not produce sufficient change in the Pc-pi c gradient to elevate edemagenesis when pulmonary capillary pressure is limited to 22 mm Hg in dogs.

Preparation and high-performance size-exclusion chromatographic (HPSEC) analysis of fluorescein isothiocyanate-hydroxyethyl starch: macromolecular probes of the blood-lymph barrier.

A procedure is described for the preparation of fluorescein isothiocyanate-labeled hydroxyethyl starch (FITC-HES). Chromatographic techniques for the purification and analysis of FITC-HES that include the development of a high-performance size-exclusion chromatographic (HPSEC) technique with a fluorescence spectrophotometer-computer detection system are described. FITC-HES macromolecules have a wide range of hydrodynamic radii (ae greater than 120- less than 10 A) and the substitution ratio of FITC to the size-selected HES macromolecules remains constant throughout the chromatographic range. The predominant isoelectric point (pI) of the multiple acidic isomers of FITC-HES is 4.6. In vitro, the very large FITC-HES macromolecules (greater than 100 A) are rapidly degraded (15 sec) by alpha-amylase in control dog plasma. While most of the large molecules (100-20 A) remain intact for greater than 24 hr, this degradation is not associated with the loss of FITC from HES. In vivo, the rate of this reaction appeared to be accelerated and the degradation of FITC-HES between 0.1 and 12 hr was small. Illustration of the HPSEC quantitation of the spectrum of FITC-HES macromolecules in "near steady-state" lymph (L) and plasma (P) samples and the L/P ratio show that this technique can be used to describe the size selectivity of the blood-lymph barrier under conditions of unaltered capillary pressure. We propose that the size-selected solvent-drag reflection coefficient (sigma f) curves for the anionic FITC-HES under conditions of elevated capillary pressure is a measure of macromolecular convective permeability of the blood-lymph barrier.

Disseminated intravascular coagulation following Echis carinatus venom in dogs: effects of a synthetic thrombin inhibitor.

Disseminated intravascular coagulation (DIC) was produced by an infusion of a prothrombin activator (Echis carinatus venom; 30 minutes; 0.5 NIH thrombin equivalent U/kg) in mongrel dogs (Echis group, n = 7). Fibrinogen declined to below measurable levels (less than 25 mg/dl), and fibrin-fibrinogen degradation products appeared (53 +/- 8 micrograms/ml) at end venom infusion in the Echis group. These alterations were not seen when an irreversible thrombin inhibitor, D-phenylalanyl-L-prolyl-L-arginine-L-chloromethyl ketone (PPACK) (57 nmol/kg/min for 120 minutes), was given alone (PPACK group, n = 5) or in association with venom (Echis + PPACK group, n = 5). Factor II activity (1% +/- 1%) in the Echis and Echis + PPACK groups was significantly below the PPACK (55% +/- 9%) and the control (79% +/- 2%) levels at 120 minutes. In contrast, factor VIII coagulant (factor VIII:C) activity in the Echis group (1% +/- 1%) remained significantly below that in the Echis + PPACK (68% +/- 8%), PPACK (78% +/- 10%), and control (91% +/- 9%) groups at this interval. No change in factors X (91% +/- 7% to 81% +/- 7%, P not significant) and VII (64% +/- 10% to 48% +/- 11%, P not significant) activities were observed. Hemolysis was observed only in the Echis group, whereas thrombocytopenia and leukopenia were noted in both the Echis and the Echis + PPACK groups. These data show that large amounts of E. carinatus venom produce rapid DIC in vivo, because of the activation of prothrombin. In contrast, the decline in factor VIII:C activity appeared to be the result of the liberated thrombin. PPACK antagonized all of the venom-released thrombin without any major deleterious clotting abnormalities. This inhibitor appears to prevent thrombin-mediated DIC in vivo. In contrast, heparin was found to be an unreliable antagonist of the venom-released thrombin in vitro. PPACK also inhibited the marked hemolysis usually observed after venom. In addition, we found that the esterolytic (N-benzoyl-L-prolyl-L-phenylalanyl-L-arginine-p-nitroanilide HCL) activity of E. carinatus venom degrades fibrinogen in vitro.

Puff adder venom shock: a model of increased vascular permeability.

The circulatory, respiratory, metabolic, lethal and tissue permeability effects of an i.v. infusion (30 min; 1.5, 2.0, 2.5 and 3.0 mg/kg) of puff adder (Bitis arietans) venom were studied in Sprague-Dawley rats (n = 35, 275-325 g). Venom (2.5 mg/kg) produced circulatory failure with arterial hypotension, hemodilution, hypoproteinemia, lactacidemia and marked hyperventilation by +4 hr. In a separate test (n = 20, 282-325 g) blood volume was measured at end venom (2.5 mg/kg) infusion (0 time) and at +3 hr with radioiodinated (125I) human serum albumin and 51Cr-labeled rat red blood cells. Venom produced a significant reduction in total blood volume index (9%, P less than .05), plasma volume index (12%, P less than .01) and red cell mass index (6%, P = N.S.) as compared to the control group at 0 time. Critically low levels of these indices were observed (43, 42 and 46%, respectively) at +3 hr. At both intervals the transvascular escape rate of radioiodinated human serum albumin but not 51Cr-labeled rat red blood cells was significantly increased in comparison to the control group. Tissue permeability index to 51Cr-labeled rat red blood cells and radioiodinated human serum albumin was increased primarily in the stomach and small intestine. These data suggest that increased vascular permeability to protein and red cells, primarily in the splanchnic region, leads to hypovolemic shock and death after a lethal dose of i.v. Bitis venom in rats.

Pulmonary fibrin microembolism with Echis carinatus venom in dogs: effects of a synthetic thrombin inhibitor.

We produced pulmonary fibrin microembolism using an infusion of a prothrombin activator (Echis carinatus venom, 30 min, 0.5 NIH thrombin equivalent units/kg) in open-chest mongrel dogs. To determine the nonclotting effects of this venom on edemagenesis we infused an irreversible thrombin inhibitor, D-phenylalanyl-L-prolyl-L-arginine chloromethyl ketone (PPACK, 57 nmol X kg-1 X min-1 for 120 min), alone (n = 5) or with venom (Echis + PPACK, n = 5). The control group (n = 5) was given 1 ml of 0.9% NaCl. A decline in left atrial pressure (means +/- SE, 5.3 +/- 0.4 to 4.0 +/- 0.5 mmHg, P less than 0.05) and cardiac index (149 +/- 10 to 82 +/- 13 ml X min-1 X kg-1, P less than 0.01) in association with a marked increase in pulmonary arterial pressure (14.5 +/- 0.6 to 26.6 +/- 2.5 mmHg, P less than 0.001) and pulmonary vascular resistance (64 +/- 5 to 304 +/- 42 mmHg X ml-1 X min-1 X kg-1, P less than 0.001) was observed after 20 min of venom infusion. During this interval, pulmonary artery wedge pressure increased (4 +/- 1 to 12 +/- 4 mmHg, P less than 0.01) in four of eight animals. Fibrinogen declined below measurable levels and fibrin microemboli were seen in many pulmonary arterioles. These changes were not observed in the Echis + PPACK, PPACK, or control groups. Leukopenia and thrombocytopenia were observed in the Echis and Echis + PPACK groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Hypotensive and hemostatic properties of rattlesnake (Crotalus viridis helleri) venom and venom fractions in dogs.

Hypotensive and hemostatic properties of Southern Pacific rattlesnake (Crotalus viridis helleri) venom (100 micrograms/kg; i.v. bolus) or venom fractions (40 micrograms/kg) were studied in mongrel dogs (n = 27, 15-27 kg). Venom was separated by gel filtration (Sephadex G-100) into three lethal fractions (FR I, II and III). Nonreduced crude venom contained 11 main protein bands on sodium dodecyl sulfate-10% polyacrylamide gel electrophoresis with a molecular weight range of 104 to 13 kD. FR I contained nine of these prominent bands (104-27 kD) and FR II contained five bands (34-13 kD). A 19-13 kD band comprised 92% of FR III. Crude venom rapidly produced hypotension, thrombocytopenia, hemolysis, the generation of fibrin monomers and a decrease in Factor VIIIC activity. Fibrin/fibrinogen degradation products appeared at +15 min and fibrinogen concentration was significantly depressed at +120 min. There was no significant change in prothrombin time or concentration of Factors VII, X and XII activity. However, the partial thromboplastin time was increased (P less than .05) by +30 min. Our data show that a high molecular weight thrombin-like venom component in FR I (greater than 27 kD) directly digests fibrinogen without activation of extrinsic and intrinsic coagulation factors. In addition, this fraction leads to the activation of the fibrinolytic system. It appears to be likely that the thrombocytopenia is primarily the result of ca. 27 kD platelet aggregating protein found in similar amounts in FR I and II. Venom components in FR II (ca. 34 and 29 kD,) were associated with hypotension and hemolysis.(ABSTRACT TRUNCATED AT 250 WORDS)