Karl-E. Arfors

Increase in vascular permeability induced by leukotriene b4 and the role of polymorphonuclear leukocytes

Leukotriene B4 (LTB4), a metabolite of arachidonic acid, is known to be a potent chemotactic and chemokinetic substance. We have used the hamster cheek pouch microcirculation model to study the effect of LTB4 on vascular permeability and the involvement of neutrophil granulocytes in this response. Intravascular fluorescein-labeled dextran (mol wt 150,000) was used as a tracer of macromolecular permeability. Topical application of LTB4 (150 nM-5 μM) to the hamster cheek pouch resulted in an immediate increase in adhering leukocytes in postcapillary venules and later larger venules. Leukocyte accumulation was reversible, but continued longer the higher the dose of LTB4 used. Subsequently, a dose-dependent increase in vascular permeability was seen at postcapillary and larger venules, with a maximum 10–20 min after application; the maximum occurred later the higher the dose of LTB4. Depletion of neutrophil granulocytes by pretreatment of the animals with antineutrophil serum obtained from immunized rabbits significantly decreased the permeability response to LTB4, whereas the response to histamine was unaffected. These results suggest that neutrophil granulocytes play a role in LTB4-mediated permeability increase. LTB4 may be of importance both for the leukocyte accumulation and for the edema formation seen in inflammatory reactions.

Muscle blood flow disturbances produced by simultaneously elevated venous and total muscle tissue pressure

Abstract The effect of simultaneously increased mean venous ( P v ) and total intramuscular pressure ( P im ) on arteriolar diameter ( D a ) and capillary blood flow ( q c ) was investigated in rabbit tenuissimus muscle, using intravital microscopy. P v and P im were increased simultaneously by placing the hind paw of the rabbit in a sealed box and elevating the pressure in this box ( P b ) with a mixture of 5% CO 2 and 95% N 2 . The mean arterial pressure outside the box ( P a ob ), was also measured. Initial D a varied between 7.6 and 45 μm ( x = 18 μ m ). In all but one measurement D a increased when P a ob - P b was lowered. The value of P a ob - P b at which q c in the tenuissimus muscle stopped was 24.3 ± 1.26 mm Hg ( x ± SEM ). When subsequently P b was lowered, muscle q c started at a value of P a ob - P b of 30.9 ± 1.56 mm Hg ( x ± SEM ), which was significantly higher than the value at which q c stopped. These data indicate that the arterioles are not the limiting factor in the muscle blood flow disturbances due to a simultaneous increase in P im and P v as in compartmental syndromes.

Changes in macromolecular permeability by intravascular generation of oxygen-derived free radicals

Abstract It has been suggested that oxygen derivatives, namely, superoxide anion radical O . 2 , hydrogen peroxide H2O2, and hydroxyl radical OH. may play a major role in inflammatory diseases, reperfusion damage after ischemia, and radiation-induced tissue deterioration. Studies of the microcirculation, using the hamster cheek pouch preparation, were carried out in order to assess possible induction of permeability changes by these species. Oxygen-derived free radicals were generated enzymatically in the microvessels by injecting xanthine oxidase via a micropipet. Permeability changes were measured by counting the number of sites at which the systemically circulating fluorescent tracer FITC-dextran ( M w = 150,000 ) leaked from postcapillary and collecting venules. Additionally, the average light intensity was assessed by means of a photomultiplier. In the cheek pouches which had received a microinjection of xanthine oxidase (0.05 unit/ml) the number of leakage sites reached a maximum of 53.0 ± 32 (mean ± SD) after 40 min, compared to 4.7 in the control group. The photomultiplier readings showed a peak after 50 min, increasing by 30.7 ± 17% (mean ± SD) of the control value before microinjection, while the control group showed no statistically significant change. Intravenous infusion of native superoxide dismutase (SOD) prevented the permeability changes completely. The same effect was obtained with intravenous injection of polyethylene glycol (PEG)-SOD. These results may indicate that administration of PEG-SOD may be an adequate approach for preventing the occurrence of O . 2 -induced alterations in microvascular permeability in various diseases.

A titanium rabbit ear chamber: assembly, insertion and results.

Abstract The technique of assembly and insertion of a titanium rabbit ear chamber is described. The method permits high-resolution studies of intravascular cellular behaviour and microcirculatory lymphatics over extended periods in conscious animals.

Effect of oxymetazoline on nasal and sinus mucosal blood flow in the rabbit as measured with laser-Doppler flowmetry.

The effect of topical oxymetazoline hydrochloride on the blood flow of the nasal and sinus mucosa of the rabbit was measured by laser-Doppler flowmetry. Oxymetazoline, the active component in clinically used nose drops, induced a dose-dependent decrease of the nasal mucosal blood flow. This effect has previously been shown in humans and suggests the presence of α2–adrenoceptors in the nasal mucosa of the rabbit. Doses of oxymetazoline used clinically in humans induced a 50% reduction of blood flow in rabbits. Rhythmic variations in blood flow were seen in 30% of the rabbits after administration of oxymetazoline. Additionally, oxymetazoline induced a dose-dependent decrease of the mucosal blood flow in the maxillary sinus when the drug was applied in the nose. A vasoconstricting effect of oxymetazoline on the arteries penetrating the maxillary sinus ostium is a possible explanation. This can have positive as well as negative consequences on acute sinus infections.

Anaphylaxis in the Monkey: Respiratory Mechanics, Acid‐base Status and Blood Gases

Aggregate anaphylaxis was induced by intravenous injection of the specific antigen in eight ovalbumin‐sensitized monkeys. Changes in respiratory mechanics, acid‐base status and blood gases were studied during the following half hour. Within 1 minute after challenge, a short period of respiratory depression, probably reflex‐mediated, was observed. This was followed by hyperventilation, and arterial Pco2 decreased. There was a rapid increase in pulmonary resistance (Rpulm) and a concomitant decrease in pulmonary dynamic compliance (Cdyn), suggesting constriction of smooth muscles in the lung. Rpulm returned to the control value but Cdyn remained depressed, as a result of constriction of small airways and pulmonary congestion. Oxygen saturation in arterial blood decreased slightly due to a marked destruction of mixed venous blood and increased venous admixture. Progressive metabolic acidosis developed, indicating poor tissue oxygenation and perfusion. The changes observed in this study were not severe enough to cause any major disturbance of the gas exchange in the lungs, despite a severe anaphylactic shock.

Fluorescence measurements in nanolitre samples

Abstract A method for the measurement of fluorescence in nanolitre samples is described. The method is simple and rapid, and allows measurement of two fluorochromes in a mixture by using the appropriate filter combinations. The sample is placed in a haemocytometer of 0.1 mm depth filled with paraffin oil. The measurement is performed with a Leitz Ortolux microscope fitted with a photomultiplier and dark-field excitation system. A stabilised xenon high pressure lamp is used as the light source. The filter combinations used for measurement of FITC-dextran and RITC-dextran are given. The results show a linear relationship between the fluorescence and concentration over a wide range, and the coefficient of variation of the method is estimated less than 3%.

Anaphylaxis in the Monkey: Pulmonary Oedema after Pre-treatment with β-Receptor Stimulants

Aggregate anaphylaxis was induced in seven ovalbumin‐sensitized monkeys, with high titres of ovalbumin specific haemagglutinating antibodies. After pre‐treatment with an intravenous (i.v.) injection of 0.25 mg/kg terbutaline (n = 6) cr an infusion of isoprenaline (n = 1), anaphylactic shock was induced by i.v. challenge with specific antigen. Haemodynamics, regional blood flows, respiratory mechanics, blood gases and haematological changes were studied during the following 30 min. Severe shock developed following ovalbumin challenge and the cardiac output was reduced by a mean of 74%. Pulmonary vascular resistance increased 11‐fold. Pulmonary dynamic compliance decreased, but there was only a minor increase in pulmonary resistance. Hypoxaemia and severe metabolic acidosis developed. Circulating platelets and leucocytes decreased markedly. Three animals died with fulminant pulmonary oedema. In conclusion, the reaction pattern was similar to that found in studies of monkeys that received no prior treatment. However, the occurrence of pulmonary oedema suggests that the effects of large doses of terbutaline on the heart, combined with the high pulmonary vascular resistance, resulted in more severe pulmonary changes than took place in untreated animals.

In vitro determination of the force required for detachment of human polymorphonuclear granulocytes (PMN) from cultured endothelium and immobilized albumin

Detachment of adherent human PMN from cultured endothelial cells (human umbilical vein, HUVEC) and from immobilized albumin (bovine serum albumin, BSA) was investigated as a function of shear stress. Non-activated PMN did not adhere to either of the substrates «2%). 20 min after activation with PMA (phorbol myristate acetate, 10 nM), 48 ± 5% (mean ± SEM, n=15) of the total number of PMN added had adhered to albumin, and 18 ± 4% (n=9) to HUVEC. The dishes with pre-adhered PMN were subjected to shear stress in the range occurring in venules in vivo, using a rotating plate-and-cone (cone angle 2.50 ) apparatus made of transparent plastic to allow observation of the cells during the assay. On HUVEC, the majority of adhered PMN (66 ± 3%, n=9) was removed by a shear stress of 0.12 Pa (1.2 dyn/cm2), while the same shear stress removed only 15 ± 5% (n=9) of the PMN adhered to BSA. Further increase of shear stress to 2.3 Pa (23 dyn/cm2) removed 39 ± 4% (n=14) of the PMN adhered to BSA. Using spherical and semispherical approximations for the shape of the adhered PMN, the adhesive forces between PMA-activated PMN and HUVEC were calculated to be on the order of 0.05 nN per PMN, while most PMN on BSA adhered with a force of more than 4 nN per PMN. The force needed to detach PMN from HUVEC is more than one order of magnitude lower than value reported for PMN adhesion to venular endothelium in vivo. The present results suggest that the endothelium may playa regulatory role in PMN adhesion and limit the attachment force to allow emigration.