Coagulation and fibrinolysis in acute mountain sickness and beginning pulmonary edema

To examine whether intravascular coagulation and/or decreased fibrinolysis precedes high-altitude pulmonary edema (HAPE) we examined 25 male mountaineers (median age 40 yr) at low altitude (550 m) and after 6, 18, and 42 h at an altitude of 4,559 m, which was climbed in 24 h. In 14 subjects, 2 of whom showed radiological evidence of HAPE after 42 h, symptoms of acute mountain sickness (AMS) were mild or absent. Eleven subjects suffered from AMS, six of whom developed radiologically documented HAPE after 18 or 42 h. In the absence of AMS there were no significant changes at high altitude, with the exception of a decrease in bleeding time from 246 +/- 18 to 212 +/- 13 (SE) (P less than 0.05). In AMS, partial thromboplastine time decreased from 34.2 +/- 0.8 to 31.1 +/- 0.5 s (P less than 0.001) and factor VIII procoagulant activity and von Willebrand factor antigen were increased by 57 +/- 12 and 70 +/- 13%, respectively (P less than 0.001), whereas there were no significant changes in beta-thromboglobulin (BTG), fibrinopeptide A (FPA), and fibrin fragment B beta 15-42. In subjects with HAPE, BTG, FPA, and B beta 15-42 were normal before and in beginning HAPE. Preceding HAPE, euglobulin clot lysis time declined at high compared with low altitude from 289 +/- 48 to 201 +/- 42 min without venous occlusion (VO) and from 107 +/- 36 to 86 +/- 31 min after VO (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

The lung at high altitude: bronchoalveolar lavage in acute mountain sickness and pulmonary edema

High-altitude pulmonary edema (HAPE), a severe form of altitude illness that can occur in young healthy individuals, is a noncardiogenic form of edema that is associated with high concentrations of proteins and cells in bronchoalveolar lavage (BAL) fluid (Schoene et al., J. Am. Med. Assoc. 256: 63-69, 1986). We hypothesized that acute mountain sickness (AMS) in which gas exchange is impaired to a milder degree is a precursor to HAPE. We therefore performed BAL with 0.89% NaCl by fiberoptic bronchoscopy in eight subjects at 4,400 m (barometric pressure = 440 Torr) on Mt. McKinley to evaluate the cellular and biochemical responses of the lung at high altitude. The subjects included one healthy control (arterial O2 saturation = 83%), three climbers with HAPE (mean arterial O2 saturation = 55.0 +/- 5.0%), and four with AMS (arterial O2 saturation = 70.0 +/- 2.4%). Cell counts and differentials were done immediately on the BAL fluid, and the remainder was frozen for protein and biochemical analysis to be performed later. The results of this and of the earlier study mentioned above showed that the total leukocyte count (X10(5)/ml) in BAL fluid was 3.5 +/- 2.0 for HAPE, 0.9 +/- 4.0 for AMS, and 0.7 +/- 0.6 for controls, with predominantly alveolar macrophages in HAPE. The total protein concentration (mg/dl) was 616.0 +/- 3.3 for HAPE, 10.4 +/- 8.3 for AMS, and 12.0 +/- 3.4 for controls, with both large- (immunoglobulin M) and small- (albumin) molecular-weight proteins present in HAPE.(ABSTRACT TRUNCATED AT 250 WORDS)     

Echocardiographic and invasive measurements of pulmonary artery pressure correlate closely at high altitude

Exaggerated hypoxia-induced pulmonary hypertension is a hallmark of high-altitude pulmonary edema (HAPE) and plays a major role in its pathogenesis. Many studies of HAPE have estimated systolic pulmonary arterial pressure (SPAP) with Doppler echocardiography. Whereas at low altitude, Doppler echocardiographic estimation of SPAP correlates closely with its invasive measurement, no such evidence exists for estimations obtained at high altitude, where alterations of blood viscosity may invalidate the simplified Bernoulli equation. We measured SPAP by Doppler echocardiography and invasively in 14 mountaineers prone to HAPE and in 14 mountaineers resistant to this condition at 4,559 m. Mountaineers prone to HAPE had more pronounced pulmonary hypertension (57 +/- 12 and 58 +/- 10 mmHg for noninvasive and invasive determination, respectively; means +/- SD) than subjects resistant to HAPE (37 +/- 8 and 37 +/- 6 mmHg, respectively), and the values measured in the two groups as a whole covered a wide range of pulmonary arterial pressures (30-83 mmHg). Spearman test showed a highly significant correlation (r = 0.89, P < 0.0001) between estimated and invasively measured SPAP values. The mean difference between invasively measured and Doppler-estimated SPAP was 0.5 +/- 8 mmHg. At high altitude, estimation of SPAP by Doppler echocardiography is an accurate and reproducible method that correlates closely with its invasive measurement.     

Evidence supportive of impaired myocardial blood flow reserve at high altitude in subjects developing high-altitude pulmonary edema

An exaggerated increase in pulmonary arterial pressure is the hallmark of high-altitude pulmonary edema (HAPE) and is associated with endothelial dysfunction of the pulmonary vasculature. Whether the myocardial circulation is affected as well is not known. The aim of this study was, therefore, to investigate whether myocardial blood flow reserve (MBFr) is altered in mountaineers developing HAPE. Healthy mountaineers taking part in a trial of prophylactic treatment of HAPE were examined at low (490 m) and high altitude (4,559 m). MBFr was derived from low mechanical index contrast echocardiography, performed at rest and during submaximal exercise. Among 24 subjects evaluated for MBFr, 9 were HAPE-susceptible individuals on prophylactic treatment with dexamethasone or tadalafil, 6 were HAPE-susceptible individuals on placebo, and 9 persons without HAPE susceptibility served as controls. At low altitude, MBFr did not differ between groups. At high altitude, MBFr increased significantly in HAPE-susceptible individuals on treatment (from 2.2 +/- 0.8 at low to 2.9 +/- 1.0 at high altitude, P = 0.04) and in control persons (from 1.9 +/- 0.8 to 2.8 +/- 1.0, P = 0.02), but not in HAPE-susceptible individuals on placebo (2.5 +/- 0.3 and 2.0 +/- 1.3 at low and high altitude, respectively, P > 0.1). The response to high altitude was significantly different between the two groups (P = 0.01). There was a significant inverse relation between the increase in the pressure gradient across the tricuspid valve and the change in myocardial blood flow reserve. HAPE-susceptible individuals not taking prophylactic treatment exhibit a reduced MBFr compared with either treated HAPE-susceptible individuals or healthy controls at high altitude.     

Changes in plasma angiotensin-converting enzyme activity and noradrenaline responses to long-term nitric oxide inhibition vary depending on their basal values in chickens

In this study, we investigated the effects of N(omega)-nitro-L-arginine (L-NNA) on arterial blood pressure (BP), plasma noradrenaline (NA) and adrenaline (A) levels and angiotensin-converting enzyme (ACE) activity. L-NNA was applied with tap water (1 mg/ml) from the 3rd to the 8th week of age (group L-NNA1). In Experiment 1, long-term L-NNA application increased BP compared to the control group (group C1) (L-NNA1 = 131.4 +/- 6.3, n = 6; C1 = 82.7 +/- 4.7 mm Hg, n = 7) but decreased plasma noradrenaline and adrenaline levels and ACE activity (NA levels: C1 = 15.5 +/- 0.8, n = 7; L-NNA1 = 8.6 +/- 0.5 ng/ml, n = 7; A levels: C1 = 15.5 +/- 0.8, n = 7; L-NNA1 = 6.0 +/- 0.5 ng/ml, n = 7; ACE activities: C1 = 87.3 +/- 3.1, n = 6; L-NNA1 = 46.2 +/- 1.9 U/l, n = 5). On the other hand, in Experiment 2 (carried out under the same conditions and in age-matched chickens), blood pressure, plasma noradrenaline levels and ACE activity were found to differ in the control group (C2) (BP = 141.4 +/- 15.5 mm Hg, n = 7; NA = 1.1 +/- 0.4 ng/ml, n = 7; ACE = 57.2 +/- 5.3 U/l, n = 7) as compared to C1, while plasma adrenaline levels were similar. In this series, long-term L-NNA application (group L-NNA2) did not change the BP, but surprisingly increased noradrenaline and ACE values (values of L-NNA2: BP = 165.7 +/- 15.6 mm Hg, n = 7; NA = 9.3 +/- 1.3 ng/ml, n = 8; ACE = 149.4 +/- 16 U/l, n = 8) while decreasing plasma adrenaline levels. L-arginine addition to L-NNA treatment completely reversed plasma noradrenaline and ACE activity values. These results indicate the modulatory activity of an L-arginine-NO pathway on adrenaline release as well as on the renin-angiotensin system in chickens.     

Hemostasis and fibrinolysis in patients with intermittent claudication: effects of prostaglandin E1

There is evidence that the coagulation system is activated in patients with peripheral arterial occlusive disease (PAOD). The beneficial effects of the vasoactive drug prostaglandin E1 (PGE1) may rely in part on the modulation of the coagulation system. The study was designed to evaluate the effects of PGE1 on hemostatic and fibrinolytic variables in patients with intermittent claudication. Therefore molecular markers of thrombin (prothrombin fragment 1+2, PTF 1+2; thrombin-antithrombin III complexes, TAT) and fibrin formation (fibrinopeptide A, FPA) and markers of the fibrinolytic activity (fibrin degradation products, D-dimers) were determined before and immediately after the first PGE1 dose (60 microg in 100 ml NaCl over 2 h i.v.) as well as after 4 weeks of daily infusion therapy in 12 PAOD patients and in eight control patients before and after a single placebo infusion. Plasma levels of PTF1+2, TAT, FPA and D-dimers tended to decrease after the initial dose of PGE1. Infusion therapy with PGE1 for 4 weeks led to a decrease of all hemostatic and fibrinolytic parameters with most pronounced changes for PFT1+2, D-dimers and plasminogen activator inhibitor-1 decreasing by 11% (P<0.05), 20% (P<0.05), and 7% (P<0.05), respectively. These variables remained unchanged in controls with placebo infusion.In summary, infusion therapy with PGE1 in patients with PAOD reduces thrombin formation and results in a decrease of fibrin degradation. PGE1 may thus reduce fibrin deposition involved in the pathogenesis of atherosclerosis.     

Hemodynamic responses to acute hypoxia, hypobaria, and exercise in subjects susceptible to high-altitude pulmonary edema

To verify the presence of the constitutional abnormality implicated in the pathogenesis of high-altitude pulmonary edema (HAPE), we evaluated the hemodynamic responses to hypoxia, hypobaria, and exercise in HAPE-susceptible subjects (HAPE-S). HAPE-S were five males with a history of HAPE. Five healthy volunteers who had repeated experiences of mountain climbing without any history of altitude-related problems served as controls. HAPE-S showed much greater increase in pulmonary vascular resistance index (PVRI) than the control subjects, resulting in a much higher level of pulmonary arterial pressure (Ppa), under both acute hypoxia of 15% O2 (Ppa = 29.0 +/- 2.8 vs. 17.8 +/- 0.3 Torr, P less than 0.05) and acute hypobaria of 515 Torr (32.3 +/- 2.8 vs. 19.1 +/- 0.8 Torr, P less than 0.05). Also, PVRI in HAPE-S exhibited a tendency to increase even during light exercise with supine bicycle ergometer (50 W), whereas PVRI in the control subjects significantly decreased, so that HAPE-S showed a greater increase in Ppa (delta Ppa = 16.0 +/- 1.5 vs. 4.9 +/- 1.1 Torr, P less than 0.001) and a greater decrease in arterial oxygen tension (17.8 +/- 4.7 vs. 5.6 +/- 1.7 Torr, P less than 0.05). We thus conclude that HAPE-S have a constitutional abnormality, which can be evaluated at low altitude, in the pulmonary circulatory responses to possible causative factors of HAPE such as hypoxia, hypobaria, and exercise.     

Formation of soluble fibrin oligomers in purified systems and in plasma.

The kinetic parameters for release of fibrinopeptide A (FPA) from human fibrinogen by thrombin are: Km = 2.3 X 10(-6)M and Vmax. = 1.1 X 10(-10)mol of FPA/s per unit of thrombin; for fibrin formation, Km is similar to that for FPA release, but, the conditions of the present study, Vmax. was approximately half of that for FPA release. The formation of fibrin polymer before the sol-gel transition was studied by gel-permeation chromatography combined with effluent analysis for fibrinogen antigen and residual FPA. Polymer formation in purified fibrinogen incubated with thrombin proceeded as a bimolecular association of exposed sites in a manner predicted by probability calculations and assuming random FPA cleavage. Each oligomer consisted of n molecules of fibrin monomer and two fibrinogen molecules, each of the latter lacking one FPA molecule, i.e. each oligomer, regardless of molecular size, retains two FPA molecules. The addition of 5 mM-CaCl2 to the reaction mixture changed the rate of polymer formation, so that dimer was no longer the prevalent oligomer; in the presence of Ca2+, the trimer was the oligomer in highest concentration. The polymers formed in the presence of calcium were similar in composition to those without, i.e. 2 mol of FPA/mol of oligomer. EDTA-treated plasma samples incubated for short periods of time, 30s or less, with thrombin ranging in concentration up to 1 N.I.H. unit/ml did not form clots during the 10-15 min period of observation until they were applied to the column, though a large proportion of the available FPA was cleaved (maximum 45%). The soluble polymers in plasma were mostly of the high-Mr variety (tetramer and greater); these high-Mr polymers contained less than 2 mol of FPA/mol of polymer, whereas dimer and trimer in plasma were similar to those in the purified systems, i.e. 2 mol of FPA/mol.     

Abnormal control of ventilation in high-altitude pulmonary edema

We wished to determine the role of hypoxic chemosensitivity in high-altitude pulmonary edema (HAPE) by studying persons when ill and upon recovery. We studied seven males with HAPE and seventeen controls at 4,400 m on Mt. McKinley. We measured ventilatory responses to both O2 breathing and progressive poikilocapnic hypoxia. Hypoxic ventilatory response (HVR) was described by the slope relating minute ventilation to percent arterial O2 saturation (delta VE/delta SaO2%). HAPE subjects were quite hypoxemic (SaO2% 59 +/- 6 vs. 85 +/- 1, P less than 0.01) and showed a high-frequency, low-tidal-volume pattern of breathing. O2 decreased ventilation in controls (-20%, P less than 0.01) but not in HAPE subjects. The HAPE group had low HVR values (0.15 +/- 0.07 vs. 0.54 +/- 0.08, P less than 0.01), although six controls had values in the same range. The three HAPE subjects with the lowest HVR values were the most hypoxemic and had a paradoxical increase in ventilation when breathing O2. We conclude that a low HVR plays a permissive rather than causative role in the pathogenesis of HAPE and that the combination of extreme hypoxemia and low HVR may result in hypoxic depression of ventilation.     

Effects of arterial pressure on lung capillary pressure and edema after microembolism

The effect of increased arterial pressure (Pa) on microvessel pressure (Pc) and edema following microvascular obstruction (100-micron glass spheres) was examined in the isolated ventilated dog lung lobe pump perfused with blood. Lobar vascular resistance (PVR) increased 2- to 10-fold following emboli when either Pa or flow was held constant. Microbead obstruction increased the ratio of precapillary to total PVR from 0.60 +/- 0.05 to 0.84 +/- 0.02 (SE) or to 0.75 +/- 0.06 (n = 6), as determined by the venous occlusion and the isogravimetric capillary pressure techniques, respectively. Isogravimetric Pc (5.0 +/- 0.7) did not differ from Pc obtained by venous occlusion (3.8 +/- 0.2 Torr, n = 6). After embolism, Pc in constant Pa decreased from 6.2 +/- 0.3 to 4.4 +/- 0.3 Torr (n = 16). In the constant-flow group, embolism doubled Pa while Pc increased only 40% (6.7 +/- 0.6 to 9.2 +/- 1.4 Torr, n = 6) with no greater edema formation than in the constant Pa groups. These data indicate poor transmission of Pa to filtering capillaries. Microembolism, even when accompanied by elevated Pa and increased flow velocity of anticoagulated blood of low leukocyte and platelet counts, caused little edema. Our results suggest that mechanical effects alone of lung microvascular obstruction cause minimal pulmonary edema.     

Leukotrienes increase levels of prostanoids in cerebrospinal fluid in piglets

We investigated effects of exogenous leukotrienes (C4, D4, or E4) on levels of prostanoids in cerebrospinal fluid in newborn pigs (1-5 days). A "closed" cranial window was placed over the parietal cortex. Pial arterial diameter was measured with a microscope and electronic micrometer system. Levels in cerebrospinal fluid (CSF) of 6-keto-Prostaglandin F1 alpha (6-keto-PGF1 alpha), Thromboxane B2 (TXB2), and Prostaglandin E2 (PGE2) were measured by radioimmunoassay. Topical application of leukotrienes C4, D4, or E4 (5,000 ng/ml) similarly constricted pial arteries by 15 +/- 2% (n = 14) (mean +/- SEM). In addition, leukotrienes increased levels of 6-keto-PGF1 alpha from 806 +/- 136 to 1,612 +/- 304 pg/ml (n = 13), TXB2 from 161 +/- 31 to 392 +/- 81 pg/ml (n = 10), and PGE2 from 2,271 +/- 342 to 4,636 +/- 740 pg/ml (n = 13). Each type of leukotriene had similar effects on prostanoid synthesis. In other experiments (n = 5), we found that 2.0 ng/ml PGE2 in CSF dilated pial arteries by 24 +/- 8% and that 1.0 ng/ml PGI2 dilated pial arteries by 15 +/- 6%. These results indicate that leukotrienes are able to increase levels of prostanoids in cerebral cortex.     

Effects of calcitonin gene-related peptide on renal blood flow in the rat

The effects of alpha-rat calcitonin gene-related peptide (alpha-rCGRP) on systemic and renal hemodynamics and on renal electrolyte excretion were examined in normal anesthetized rats. In one group of rats (n = 7), infusions of alpha-rCGRP at doses of 10, 50, 100, and 500 ng/kg/min for 15 min each produced dose-related and significant decreases in mean arterial pressure from a control of 130 +/- 3 mm Hg to a maximal depressor response of 91 +/- 2 mm Hg. During the first three doses of alpha-rCGRP, renal blood flow progressively and significantly increased from a control of 5.0 +/- 0.3 ml/min to a peak level of 6.3 +/- 0.3 ml/min achieved during the 100 ng/kg/min infusion. With the highest infusion rate of 500 ng/kg/min, renal blood flow fell below the control level to 4.5 +/- 0.2 ml/min (P less than 0.05). The responses in renal blood flow and mean arterial pressure were associated with reductions in renal vascular resistance. After cessation of alpha-rCGRP infusions, arterial pressure, renal blood flow, and renal vascular resistance gradually returned toward the baseline values. In another group of rats (n = 9), infusion of alpha-rCGRP for 30 min at 100 ng/kg/min produced a significant reduction in urinary sodium excretion from 0.28 +/- 0.06 to 0.14 +/- 0.5 muEq/min (P less than 0.05). Urine flow and urinary potassium excretion also appeared to decrease, but the changes were not significantly different (P greater than 0.05) from their respective baselines. These results demonstrate that alpha-rCGRP is a potent and reversible hypotensive and renal vasodilatory agent in the anesthetized rat. The data also suggest that alpha-rCGRP may have significant effects on the excretory function of the kidney.     

Effect of bronchial artery blood flow on cardiopulmonary bypass-induced lung injury

Cardiovascular surgery requiring cardiopulmonary bypass (CPB) is frequently complicated by postoperative lung injury. Bronchial artery (BA) blood flow has been hypothesized to attenuate this injury. The purpose of the present study was to determine the effect of BA blood flow on CPB-induced lung injury in anesthetized pigs. In eight pigs (BA ligated) the BA was ligated, whereas in six pigs (BA patent) the BA was identified but left intact. Warm (37 degrees C) CPB was then performed in all pigs with complete occlusion of the pulmonary artery and deflated lungs to maximize lung injury. BA ligation significantly exacerbated nearly all aspects of pulmonary function beginning at 5 min post-CPB. At 25 min, BA-ligated pigs had a lower arterial Po(2) at a fraction of inspired oxygen of 1.0 (52 +/- 5 vs. 312 +/- 58 mmHg) and greater peak tracheal pressure (39 +/- 6 vs. 15 +/- 4 mmHg), pulmonary vascular resistance (11 +/- 1 vs. 6 +/- 1 mmHg x l(-1) x min), plasma TNF-alpha (1.2 +/- 0.60 vs. 0.59 +/- 0.092 ng/ml), extravascular lung water (11.7 +/- 1.2 vs. 7.7 +/- 0.5 ml/g blood-free dry weight), and pulmonary vascular protein permeability, as assessed by a decreased reflection coefficient for albumin (sigma(alb); 0.53 +/- 0.1 vs. 0.82 +/- 0.05). There was a negative correlation (R = 0.95, P < 0.001) between sigma(alb) and the 25-min plasma TNF-alpha concentration. These results suggest that a severe decrease in BA blood flow during and after warm CPB causes increased pulmonary vascular permeability, edema formation, cytokine production, and severe arterial hypoxemia secondary to intrapulmonary shunt.     

Role of the renin-angiotensin system during alterations of sodium intake in conscious mice

The present studies were performed to quantify circulating components of the renin-angiotensin-aldosterone axis and to determine the functional importance of this system during alterations in sodium intake in conscious mice. Increasing sodium intake from approximately 200 to 1,000 microeq/day significantly decreased plasma renin concentration from 472 +/- 96 to 304 +/- 83 ng ANG I. ml(-1). h(-1) (n = 5) but did not alter plasma renin activity from the low-sodium level of 7.7 +/- 1.1 ng ANG I. ml(-1). h(-1). Despite the elevated plasma renin concentration, plasma ANG II in mice on low-sodium level averaged 14 +/- 3 pg/ml and was significantly suppressed to 6 +/- 1 pg/ml by high-sodium intake (n = 7). Consistent with the modulation of ANG II, plasma aldosterone significantly decreased from 41 +/- 8 to 8 +/- 3 ng/dl when sodium intake was elevated (n = 6). In a final set of experiments, the continuous infusion of ANG II (20 ng. kg(-1). min(-1)) led to a mild salt-sensitive increase in mean arterial pressure from 108 +/- 2 to 131 +/- 2 mmHg as sodium intake was varied from low to high (n = 7). In vehicle-infused mice, mean arterial pressure was unaltered from 109 +/- 2 mmHg when sodium intake was increased (n = 6). These studies indicate that the physiological suppression of circulating ANG II may be required to maintain a constancy of arterial pressure during alterations in sodium intake in normal mice.     

Blunted hypoxic ventilatory drive in subjects susceptible to high-altitude pulmonary edema

It has been proposed that subjects susceptible to high-altitude pulmonary edema (HAPE) show exaggerated hypoxemia with relative hypoventilation during the early period of high-altitude exposure. Some previous studies suggest the relationship between the blunted hypoxic ventilatory response (HVR) and HAPE. To examine whether all the HAPE-susceptible subjects consistently show blunted HVR at low altitude, we evaluated the conventional pulmonary function test, hypoxic ventilatory response (HVR), and hypercapnic ventilatory response (HCVR) in ten lowlanders who had a previous history of HAPE and compared these results with those of eight control lowlanders who had no history of HAPE. HVR was measured by the progressive isocapnic hypoxic method and was evaluated by the slope relating minute ventilation to arterial O2 saturation (delta VE/delta SaO2). HCVR was measured by the rebreathing method of Read. All measurements were done at Matsumoto, Japan (610 m). All the HAPE-susceptible subjects showed normal values in the pulmonary function test. In HCVR, HAPE-susceptible subjects showed relatively lower S value, but there was no significant difference between the two groups (1.74 +/- 1.16 vs. 2.19 +/- 0.4, P = NS). On the other hand, HAPE-susceptible subjects showed significantly lower HVR than control subjects (-0.42 +/- 0.23 vs. -0.87 +/- 0.29, P less than 0.01). These results suggest that HAPE-susceptible subjects more frequently show low HVR at low altitude. However, values for HVR were within the normal range in 2 of 10 HAPE-susceptible subjects. It would seem therefore that low HVR alone need not be a critical factor for HAPE. This could be one of several contributing factors.     

Soluble fibrin and D-dimer at normal pregnancy and pregnancy with risk of miscarriage

ELISA for soluble fibrin (SF) quantification has been elaborated on the basis of our fibrin-specific monoclonal antibodies (mAb). Epitope for these mAb is localized in fibrin fragment Bbeta118-134. The method was used on the blood plasma of healthy pregnant women (control group) and pregnant women with the risk of fetal loss (RFL). The increased mean values of SF concentrations were observed at pregnancy with RFL as compared to the normal pregnancy at the terms from 4 to 24 weeks (17.87 +/- 3.15 mkg/ml and 9.03 +/- 1.58 mkg/ml accordingly, p < 0.05). A weak negative correlation between SF concentration and pregnancy term was found at RFL (r = -0.201, n=35), while there was no correlation between these variables in control group (r = 0.004, n=28). The mean values of SF concentration estimated by semiquantitative test (by phosphates salting out of SF) were also higher at the pregnancy with RFL as compared to the normal pregnancy. However, the absolute values of SF concentrations determined by salting out method were essentially higher than in the case of ELISA. Immunoblot analysis with mAb 2d-2a (epitope for which in fibrin molecule encompasses peptide bond Bbeta14-15), showed that the main molecular component of SF at normal pregnancy and RFL was oligomeric fibrin desAA with possible incorporation of fibrinogen and/or fibrin desA which was not stabilized by factor XIIIa. D-dimer concentrations determined in blood plasma samples of pregnant women by ELISA varied in the range of 1-224 ng/ml at the pregnancy period from 4 to 37 weeks. There was positive correlation between D-dimer concentration and pregnancy term both at normal pregnancy and pregnancy with RFL (r = 0.765, n=33 and r = 0.712, n=44 correspondingly). The mean values of D-dimer concentration at various terms of normal pregnancy and pregnancy with RFL did not vary considerably. Thus SF but not D-dimer quantification may give useful diagnostic information at the pregnancy with RFL.     

Activation of the coagulation mechanism on tumor necrosis factor-stimulated cultured endothelial cells and their extracellular matrix. The role of flow and factor IX/IXa

Infusion of tumor necrosis factor (TNF) into tumor-bearing mice led to intravascular clot formation with fibrin deposition in microvessels in the tumor bed in close association with the vessel wall, which could be prevented by active site-blocked factor IXa (IXai). This observation prompted us to examine the role of the intrinsic system in activation of the coagulation mechanism on TNF-stimulated human endothelial cell monolayers and endothelial-derived matrix during exposure to purified coagulation factors or flowing blood. Treatment of endothelial cells in intact monolayers with TNF induced expression of the procoagulant cofactor tissue factor (TF) in a dose-dependent manner, and after removal of the cells, TF was present in the matrix. TNF-treated endothelial cell monolayers exposed to blood anticoagulated with low molecular weight heparin induced activation of coagulation. Addition of IXai blocked the procoagulant response on TNF-treated endothelial cells, and consistent with this, the presence of factor IX/VIIIa enhanced endothelial TF/factor VII(a) factor X activation over a wide range of cytokine concentrations (0-600 pM). When TF-dependent factor X activation on endothelial cells was compared with preparations of subendothelium, the extracellular matrix was 10-20 times more effective. IXai blocked TF/factor VII(a) mediated activated coagulation on matrix, but only at lower concentration of TNF (less than 50 pM). Similarly, enhancement of factor Xa formation on matrix by factors IX/VIIIa was most evident at lower TNF concentrations. When anticoagulated whole blood flowing with a shear of 300 s-1 was exposed to matrices from TNF-treated endothelial cells, but not matrices from control cells, fibrinopeptide A (FPA) generation, fibrin deposition, and platelet aggregate formation were observed. FPA generation could be prevented by a blocking antibody to TF and by active site-blocked factor Xa (Xai) over a wide range of TNF concentrations (0-600 pM), whereas IXai only blocked FPA generation at lower TNF concentrations (less than 50 pM). Activation of coagulation on matrix from TNF-stimulated endothelial cells was dependent on the presence of platelets, indicating the important role of platelets in propagating the reactions leading to fibrin formation. These observations demonstrate the potential of cytokine-stimulated endothelium and their matrix to activate coagulation and suggest the importance of the intrinsic system in factor Xa formation on cellular surfaces.     

Circulating vasoactive substances and hemodynamic adjustments at birth in lambs

Circulating vasoactive substances and hemodynamics were examined in chronically instrumented unanesthetized lambs before, during, and after cesarean section (spontaneous respiration). One of three infusions were started 20 min before birth: saline control (n = 10), saralasin (n = 5), or captopril (n = 6). Control lambs exhibited peak (means +/- SE) increases above fetal base line at 5 min after birth in plasma renin activity (5.0 +/- 1.1 to 11.0 +/- 3.4 ng.ml-1.h-1), angiotensin II (ANG II, 37 +/- 6 to 141 +/- 45 pg/ml) and total catecholamines (318 +/- 35 to 3,821 +/- 580 pg/ml). Mean systemic arterial pressure (Psa) and arterial O2 partial pressure (PaO2) increased more rapidly and to a greater extent by 1 h after birth in control lambs (Psa, 65 +/- 1 Torr; PaO2, 45 +/- 3 Torr) compared with the captopril group (Psa, 53 +/- 2 Torr; PaO2, 31 +/- 4 Torr) and the saralasin group (Psa, 56 +/- 2 Torr; PaO2, 27 +/- 3 Torr). Intravenous infusions of ANG II in control lambs, 2 h after birth resulted in a preferential systemic vs. pulmonary pressor response. The results demonstrate that at birth ANG II formation fosters the postnatal rise in Psa and PaO2, and high levels of circulating catecholamines may support postnatal cardiac output and Psa.     

Relevance of vasoactive mediators for the blood pressure effects of intravenous arachidonic acid injection in rats.

Vasopressor response and release of eicosanoids following intravenous injection of arachidonic acid (AA) were examined in normotensive rats. AA administration caused a rapid initial fall of arterial pressure followed by a brief rise and a subsequent prolonged fall in anesthetized rats. Immediately after AA injection the blood levels of TXB2 and 6-keto-PGF1 alpha, the stable metabolites of TXA2 and prostacyclin, rose, from 1.52 +/- 0.23 ng/ml to 176.4 +/- 42.6 ng/ml and from 4.05 +/- 0.67 ng/ml to 171.4 +/- 31.2 ng/ml, respectively. Blood pressure behaviour and eicosanoid blood level were influenced by different inhibitors and antagonists of vasoactive mediators. The cyclooxygenase inhibitor acetylsalicylic acid completely eliminated the second blood pressure depression after AA injection and simultaneously diminished TXB2 and 6-keto-PGF1 alpha formation in murine blood, whereas the TXA2 receptor antagonist BM 13.177 prevented the return of the blood pressure to preinjection level after the initial brief fall in arterial pressure. Although the TXA2 synthase inhibitor HOE 944 markedly inhibited TXB2 formation, no influence on AA-induced blood pressure changes could be registered. The receptor antagonist of platelet activating factor BN 52021 and the serotonin and histamine receptor antagonist cyproheptadine also reduced TXB2 amounts, in murine blood without any effects on blood pressure behaviour.     

Splenic denervation worsens lipopolysaccharide-induced hypotension, hemoconcentration, and hypovolemia

During lipopolysaccharide (LPS)-induced endotoxemia, increased intrasplenic fluid efflux contributes to a reduction in plasma volume. We hypothesized that splenic sympathetic nerve activity (SSNA), which increases during endotoxemia, limits intrasplenic fluid efflux. We reasoned that splenic denervation would exaggerate LPS-induced intrasplenic fluid efflux and worsen the hypotension, hemoconcentration, and hypovolemia. A nonlethal dose of LPS (150 microg x kg(-1) x h(-1) for 18 h) was infused into conscious male rats bearing transit time flow probes on the splenic artery and vein. Fluid efflux was estimated from the difference in splenic arterial inflow and venous outflow (A-V). LPS significantly increased the (A-V) flow differential (fluid efflux) in intact rats (saline -0.01 +/- 0.02 ml/min, n = 8 vs. LPS +0.21 +/- 0.06 ml/min, n = 8); this was exaggerated in splenic denervated rats (saline -0.03 +/- 0.01 ml/min, n = 7 vs. LPS +0.41 +/- 0.08 ml/min, n = 8). Splenic denervation also exacerbated the LPS-induced hypotension, hemoconcentration, and hypovolemia (peak fall in mean arterial pressure: denervated 19 +/- 3 mmHg, n = 10 vs. intact 12 +/- 1 mmHg, n = 8; peak rise in hematocrit: denervated 6.7 +/- 0.3%, n = 8 vs. intact 5.0 +/- 0.3%, n = 8; decrease in plasma volume at 90-min post-LPS infusion: denervated 1.08 +/- 0.15 ml/100 g body wt, n = 7 vs. intact 0.54 +/- 0.08 ml/100 g body wt, n = 8). The exaggerated LPS-induced hypovolemia associated with splenic denervation was mirrored in the rise in plasma renin activity (90 min post-LPS: denervated 11.5 +/- 0.8 ng x ml(-1) x h(-1), n = 9 vs. intact 6.6 +/- 0.7 ng x ml(-1) x h(-1), n = 8). These results are consistent with our proposal that SSNA normally limits LPS-induced intrasplenic fluid efflux.