Chemotactic factor-specific desensitization of skin to infiltration by polymorphonuclear leukocytes.

The accumulation of polymorphonuclear leukocytes (PMNs) was measured in rabbits following single or repeated injections into skin sites of the inflammatory agents, zymosan-activated plasma and formylmethionyl-leucyl-phenylalanine. Fewer cells entered lesions restimulated with the same agent during the subsequent inflammatory response than simultaneously entered skin sites stimulated for the first time. Decreased reactivity of a restimulated site was specific to the initiating stimulus, developed within 2 to 4 h of initial stimulation and persisted for at least 8 h. The results indicate that PMN accumulation in acute inflammatory lesions is regulated locally by a chemotactic factor-specific mechanism which differs from PMN deactivation and concentration gradient mechanisms currently thought to regulate PMN migration in vivo.

Plasminogen activator and thromboplastin activity from sheep alveolar macrophages.

Alveolar lavage cells from normal sheep were found to be composed of over 95% macrophages. When the cells were cultured, fibrinolytic and thromboplastin-like activities could be detected within 2-4 hours of incubation. As the number of cultured cells was increased the two activities in the conditioned medium increased proportionately. The cells were separated into two distinct subpopulations by means of a sedimentation velocity cell fractionation technique. The macrophage subpopulations were examined for differences in size, morphology, esterase staining and ability to release plasminogen activator and procoagulant activity respectively. These activities were confined to the large cell subpopulation. The fibrinolytic activity was shown to be plasminogen-dependent and could be inhibited by DFP. On the basis of this the fibrinolytic activity has been designated as plasminogen activator. The procoagulant activity was shown to be thromboplastin in nature because it was Factor VII dependent, inactivated by phospholipase C and not inhibited by DFP. The procoagulant activity has been designated as macrophage thromboplastin. The two activities could be distinguished on the basis of DFP inhibition.

Effect of a fibrin(ogen)-derived vasoactive peptide on polymorphonuclear leukocyte emigration.

A vasoactive peptide known to increase vascular permeability and corresponding to residues 30-43 of the human fibrinogen B beta-chain induced polymorphonuclear leukocyte emigration in rabbit skin in vivo. The leukocyte emigration was much stronger after 2 h than after 0.5 h. Addition of prostaglandin E2 (PGE2) did not influence the chemotactic activity, which might possibly be explained by a known PGI2 releasing capacity of this peptide. PGE2 enhanced the leukocyte emigration due to leukotriene B4.

Emigration and accumulation of PMN-leukocytes induced by endotoxin, interleukin 1 and other chemotactic substances.

This publication describes polymorphonuclear leukocyte (PMN) emigration and accumulation, which is prerequisite for their defensive function in infected tissues. The extravasated PMNs can kill microorganisms, but in this process they also release proteolytic enzymes and other cell constituents which can alter and even injure the tissues, primarily the microcirculation. In the first part of the paper in vivo quantitation of the acute inflammatory reaction is described with emphasis on PMN emigration and accumulation. With 51Cr-labeled PMNs the kinetics of their emigration induced by a number of chemotaxins and chemotaxinigens was found to be similar, peaking in 1-4 hour old lesions and returning to baseline values thereafter. The most potent substance tested was endotoxin, which induced a PMN influx at a molar concentration a least 3 orders of magnitude lower than the other substances tested, implying the these substances are not the primary endogenous mediators of endotoxin induced inflammation. Next we describe an observation which shed considerable light on the mechanisms underlying PMN emigration. When a chemotaxin or endotoxin was injected intradermally and after varying periods of time reinjected into the same site, the PMN influx into those sites was diminished, compared to sites not previously injected, i. e. injected for the first time. This tachyphylaxis or diminished responsiveness was attributed to a downregulation of receptors, presumably on endothelial cells, coupled to a facilitatory mechanism. Other mechanism proposed to terminate emigration of PMNs during inflammatory reaction were unlikely, based on our experimental findings. Endotoxin is not chemotactic in vitro but it induces PMN emigration when injected intradermally. Hence the third part of the publication deals with PMN emigration induced by interleukin 1 and its significance for endotoxin-induced inflammation. IL 1 is the only chemotaxin which induces PMN accumulation at a concentration comparable to that of endotoxin and considerably lower than the other chemotaxins. There was cross tachyphylaxis between endotoxin and IL 1 and vice versa. The PMN influx into IL 1 sites injected 6 hours earlier with IL 1 or with endotoxin was diminished compared to IL 1 sites injected into normal skin. Sites injected first with IL 1 and then with a low dose of endotoxin also exhibited cross tachyphylaxis. FMLP or LTB4 injected into sites pretreated with endotoxin did not exhibit cross tachyphylaxis, i. e. the PMN influx was similar to sites injected for the first time with these chemotaxins.(ABSTRACT TRUNCATED AT 400 WORDS)

The role of histamine and other mediators in microvascular changes in acute inflammation.

In addition to bronchial smooth muscle, histamine and other mediators act in bronchial asthma on the microcirculation of pulmonary connective tissue. The mediators induce enhanced blood flow, and by acting on the blood-tissue barrier, they induce increase in vasopermeability with edema and in more severe injury microhemorrhage and microthrombosis and infiltration of the connective tissue by leukocytes, predominantly neutrophils. The scheme proposed 10 years ago by K. F. Austen and co-workers in 1976 still holds true: a short-acting humoral-cellular phase is followed by a longer-acting pathopharmacologic or inflammatory phase. Some mediators, including histamine, serotonin, bradykinin, the sulfido leukotrienes and platelet activating factor, have a direct effect on endothelium and smooth muscle cells and more severe injury is due to mediators that exert their effect via neutrophils, which release lysosomal constituents. The intact complement-derived fragments C3a and C5a act directly and for a short period, as do histamine and the other direct-action mediators. The accumulation of neutrophils is brought about by C5a and its stable derivative C5ades Arg, leukotriene B4, PAF, and interleukin-1. Unlike the direct-action mediators, whose effect on the microcirculation does not extend beyond 20-25 min, those mediating via neutrophils have a delayed effect (peak 1-2 h), which parallels the neutrophil influx. The direct action mediators exert their effects on the microcirculation also in neutropenic animals. The most potent agents causing enhanced blood flow are the prostaglandins of the E class. Through the enhanced blood flow increase in vasopermeability, neutrophil emigration and hemorrhage are enhanced.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental bacterial pneumonia in rabbits: polymorphonuclear leukocyte margination and sequestration in rabbit lungs and quantitation and kinetics of 51Cr-labeled polymorphonuclear leukocytes in E. coli-induced lung lesions.

A relationship between the circulating and marginal polymorphonuclear leukocyte (PMN) pools was documented using 51Cr-labeled leukocytes as a marker. 51Cr-leukocytes marginating in the lungs were found to decrease following a first-order exponential decline, while 51Cr radioactivity accumulated in the liver and the spleen. Intravenously administered endotoxin caused a rapid selective disappearance of PMNs from the circulation. The percentage of infused 51Cr cells disappearing was equal to the percentage of disappearance of host cells. The PMNs were found to sequester in the lungs, with peak sequestration of labeled cells occurring 5 min after an endotoxin challenge. Over the next 25 min the 51Cr radioactivity in the lungs declined. Large numbers of PMNs, probably newly derived from the bone marrow, were observed histologically to be sequestered in the lung vasculature 90 min after an endotoxin dose, while the early sequestration of circulating leukocytes could not be assessed histologically. Pulmonary inflammatory lesions were induced selectively with Escherichia coli in the left lower lobes of rabbits, leaving the right lower lobes as intrinsic controls. PMN-accumulation into the lesions was quantitated using 51Cr-labeled blood leukocytes. With the aid of 125I-labeled E. coli, a logarithmic dose-response relationship was found between the number of E. coli and of PMNs. Over a 6-hr period circulating PMNs were found to accumulate in a lesion in the left lower lobe, whereas in the control right lower lobe, leukocyte radioactivity declined. These findings were confirmed with the aid of lavages of the right and left lungs. Two peaks of PMN-accumulation were found by studying leukocyte kinetics: a larger peak between 0 and 6 hr and a smaller peak 18-24 hr after instillation of the microorganisms. Histologic studies confirmed the accumulation of leukocytes, and by 3 weeks showed a complete resolution of the lesions.

The effect of vasodilator prostaglandins on polymorphonuclear leukocyte infiltration and vascular injury.

Some severe acute inflammatory reactions are characterized by polymorphonuclear leukocyte (PMN) infiltration as well as vascular and tissue damage with hemorrhage. Two types of mediators that may be involved in such reactions are chemotactic factors and prostaglandins. The chemotactic factors can induce PMN infiltration, while some types of prostaglandins cause vasodilatation. We reported previously that injection of soluble, nonphagocytosable chemotactic stimuli, zymosan-activated plasma (ZAP), or C5a des Arg into rabbit skin induced PMN-dependent hemorrhage. Here we investigated whether prostaglandins may modulate the rate of PMN infiltration, measured with 51Cr-labeled leukocytes and the degree of hemorrhage, measured with 59Fe-labeled red cells. Prostaglandin (PG) E1 (0.5 microgram) or E2 (1 microgram) increased ZAP-induced PMN accumulation by 81% and hemorrhage by 400%. A similar potentiation by PGE2 was observed when submaximal concentrations of ZAP were injected. Prostaglandin F2 alpha had no such effect. These results indicate that the degree of PMN infiltration of the tissues may be one factor determining the severity of vascular damage. Furthermore, vasodilatory prostaglandins, generated during neutrophilic inflammatory reactions, may enhance chemotactic-factor-mediated PMN infiltration and increase the extent of vascular injury.

Kinetics of acute inflammation induced by Escherichia coli in rabbits. II. The effect of hyperimmunization, complement depletion, and depletion of leukocytes.

The inflammatory response to Escherichia coli was quantitated in the skin of normal rabbits and the kinetics established as described previously. Hyperemia, measured with radiolabeled microspheres; vascular permeability, estimated with 125 I-albumin; and leukocyte infiltration, quantitated with 51Cr-labeled autologous leukocytes, reached maximal values 3 hours after the injection of bacteria and subsided almost completely by 6 hours. Hemorrhage, measured with homologous 59Fe-erythrocytes, continued to increase between 1 and 6 hours after injection and then reached plateau levels. The lesions were studied up to 8 hours, since in the previous study no changes were observed beyond that time. In the study described in this paper, the host mediation systems were manipulated in various groups of rabbits in order to elucidate the mechanisms underlying the development of the inflammatory reaction. One group of animals was hyperimmunized with the E coli organisms, another was partially depleted of hemolytic complement with cobra venom factor, and yet another was rendered leukopenic with nitrogen mustard. In hyperimmunized animals hyperemia in the dermal lesions induced by the microorganisms was significantly more intense than in normal rabbits. Vascular permeability increase occurred earlier in hyperimmunized rabbits and at 1 hour was significantly greater than in normals. Decomplemented rabbits had significantly less vascular permeability than normal animals, whereas in leukopenic rabbits no increase in vascular permeability could be elicited. Leukocyte accumulation was increased over the normal animals in the lesions of hyperimmunized rabbits. Hemorrhage was significantly decreased in leukopenic rabbits. Histologic examination of the lesions revealed that whereas in normal animals the infiltrating neutrophils ingested most of the bacteria and formed definite abscesses by 6-8 hours, these abscesses were absent in leukopenic animals, and free-lying bacteria were demonstrable in lesions. Histologically more neutrophils were present in the hyperimmunized than in the normal rabbits, but this difference was striking when normal animals were compared with leukopenic animals, in some of which only very occasional small accumulations of neutrophils were present.