Cell surface levels of endothelial ICAM‐1 influence the transcellular or paracellular T‐cell diapedesis across the blood–brain barrier

The extravasation of CD4⁺ effector/memory T cells (T_EM cells) across the blood–brain barrier (BBB) is a crucial step in the pathogenesis of experimental autoimmune encephalomyelitis (EAE) or multiple sclerosis (MS). Endothelial ICAM‐1 and ICAM‐2 are essential for CD4⁺ T_EM cell crawling on the BBB prior to diapedesis. Here, we investigated the influence of cell surface levels of endothelial ICAM‐1 in determining the cellular route of CD4⁺ T_EM‐cell diapedesis across cytokine treated primary mouse BBB endothelial cells under physiological flow. Inflammatory conditions, inducing high levels of endothelial ICAM‐1, promoted rapid initiation of transcellular diapedesis of CD4⁺ T cells across the BBB, while intermediate levels of endothelial ICAM‐1 favored paracellular CD4⁺ T‐cell diapedesis. Importantly, the route of T‐cell diapedesis across the BBB was independent of loss of BBB barrier properties. Unexpectedly, a low number of CD4⁺ T_EM cells was found to cross the inflamed BBB in the absence of endothelial ICAM‐1 and ICAM‐2 via an obviously alternatively regulated transcellular pathway. In vivo, this translated to the development of ameliorated EAE in ICAM‐1^null//ICAM‐2^?/?C57BL/6J mice. Taken together, our study demonstrates that cell surface levels of endothelial ICAM‐1 rather than the inflammatory stimulus or BBB integrity influence the pathway of T‐cell diapedesis across the BBB.     

Hemorrhagic endovasculitis and hemorrhagic villitis of the placenta

A unique vascular lesion has been identified in 32 placentas sent to the Michigan Placental Tissue Registry, East Lansing, Mich. Half of the associated infants are stillborn; 11 of 16 liveborn were in distress or small for gestational age. Placental vessels show thrombosis, endothelial, and medial hyperplasia and narrowing or obliteration of the lumen. A microangiopathic process is suggested by intraluminal fragmentation of erythrocytes with diapedesis of intact and fragmented RBCs through vessel walls. Red blood cell fragments and hemosiderin are present within villous stroma. Nucleated erythrocytes in placental vessels suggest fetal hypoxia. On gross examination, the placentas are meconium stained. The umbilical cords are frequently edematous, redundant, and around a fetal part; gradual narrowing of placental vessels may be related. Chronic villitis is present in 75% of the cases, intranuclear inclusions have been identified in 10%. An infectious agent, possibly viral, is suggested; toxic and chemical substances must be considered.     

The endothelium in early experimental contact hypersensitivity of the oral mucosa. Leukocyte-binding properties and ultrastructure

Contact hypersensitivity is characterized by an early and specific diapedesis of mononuclear cells into the site of antigenic challenge. In order to study the functional and ultrastructural properties of the endothelium involved in the recruitment of leukocytes, Sprague-Dawley rats were skin sensitized to DNFB; and this was followed by challenge of the oral mucosa. In vitro binding of peripheral blood mononuclear cells to high endothelial venules in lymph nodes was highly specific but no affinity of peripheral blood mononuclear cells to the vessels was observed in normal oral mucosa or in early contact hypersensitivity. However, 10 days after repeated DNFB challenge, occasional vessels bound overlaid peripheral blood mononuclear cells. Ultrastructurally, we identified migration of mononuclear cells through small venules three h after challenge. The vessels involved, however, did not display morphological signs of activation reminiscent of high endothelial venules in lymph nodes. Mast cell degranulation was evident as early as 30 min after challenge, and a possible mechanism for mast cell-mediated leukocyte recruitment is discussed.     

Ultrastructural evidence for extravascular platelet recruitment in the lung upon intravenous injection of platelet-activating factor (PAF-acether) to guinea-pigs

Bronchoconstriction and degenerative lesions of the bronchial epithelium were observed microscopically 1 min after the i.v. injection of 100 ng/kg of platelet-activating factor (PAF-acether) to the anaesthesized guinea-pig. Constricted arterioles containing marginated polymorphonuclear neutrophils and platelet aggregates were seen, as well as alveolar capillaries obstructed by thrombi formed by partially or totally degranulated platelets. Three minutes after the injection of PAF-acether, platelet diapedesis to the alveolar septa and lumen was clearly observed. Bronchoconstriction was still present at 3 min, but subsided after 60 min, whereas oedema of the submucosa persisted accompanied by an infiltration of eosinophils and neutrophils. The infusion of prostacyclin prevented the formation of platelet aggregates and platelet diapedesis due to PAF-acether, but the morphological evidence of bronchial constriction was not modified. Aspirin failed completely to modify the effects of PAF-acether. Our results show that PAF-acether causes early formation and deposition of platelet aggregates, accompanied by the margination of polymorphonuclear neutrophil leucocytes in pulmonary vessels of the guinea-pig. Since bronchoconstriction persisted when platelet aggregation was inhibited with prostacyclin, aggregation by itself would not account for this effect. Early platelet diapedesis in the vicinity of bronchial smooth muscle corroborates previous evidence that platelets contain and release bronchoconstrictor substances, which operate by cyclo-oxygenase-independent mechanisms and are possibly involved with the physiopathology of lung inflammation during immediate hypersensitivity.     

Ultrastructural evidence for extravascular platelet recruitment in the lung upon intravenous injection of platelet-activating factor (PAF-acether) to guinea-pigs.

Bronchoconstriction and degenerative lesions of the bronchial epithelium were observed microscopically 1 min after the i.v. injection of 100 ng/kg of platelet-activating factor (PAF-acether) to the anaesthesized guinea-pig. Constricted arterioles containing marginated polymorphonuclear neutrophils and platelet aggregates were seen, as well as alveolar capillaries obstructed by thrombi formed by partially or totally degranulated platelets. Three minutes after the injection of PAF-acether, platelet diapedesis to the alveolar septa and lumen was clearly observed. Bronchoconstriction was still present at 3 min, but subsided after 60 min, whereas oedema of the submucosa persisted accompanied by an infiltration of eosinophils and neutrophils. The infusion of prostacyclin prevented the formation of platelet aggregates and platelet diapedesis due to PAF-acether, but the morphological evidence of bronchial constriction was not modified. Aspirin failed completely to modify the effects of PAF-acether. Our results show that PAF-acether causes early formation and deposition of platelet aggregates, accompanied by the margination of polymorphonuclear neutrophil leucocytes in pulmonary vessels of the guinea-pig. Since bronchoconstriction persisted when platelet aggregation was inhibited with prostacyclin, aggregation by itself would not account for this effect. Early platelet diapedesis in the vicinity of bronchial smooth muscle corroborates previous evidence that platelets contain and release bronchoconstrictor substances, which operate by cyclo-oxygenase-independent mechanisms and are possibly involved with the physiopathology of lung inflammation during immediate hypersensitivity.     

Leukemic cell-endothelial cell interactions in leukemic cell dissemination

In some human and experimental leukemias and lymphomas, the pattern of metastasis can be correlated with the homing sites of the normal progenitor cells. In vitro binding assays and homing experiments with murine lymphoma cell lines suggest that the nonrandom distribution of metastasis could be determined by specificity of cell-endothelium binding. A single subcutaneous inoculum of L2C cells in strain II guinea pigs resulted in a predictable stereotyped pattern of metastasis. Leukemic cell infiltrates were mainly observed around veins at specific locations in each organ: in brain, the most superficial leptomeningeal veins; in liver, veins of the portal triads; in lung, peribronchial veins; in kidney, veins of the renal columns; in adrenal gland, capsular veins and veins of the medulla. Those vessels also showed intense leukemic cell binding and diapedesis. This would suggest that the leukemic infiltrates were the result of transvenular traffic in these regions. Because leukemic cells were both in the intra- and extravascular compartments, the direction of the cell migration could not be determined. When L2C cells were injected into the right auricle of normal guinea pigs, leukemic cell binding occurred almost exclusively in veins located in areas of metastasis predilection. In addition, extravasation by diapedesis occurred in high endothelial venules in lymphoid organs, peribronchial veins and veins in the portal triads. Neither leukemic cell binding nor diapedesis occurred at the sinusoidal or capillary levels; extravasation in these vessels results from intravascular proliferation and secondary damage of the vessel wall, not by diapedesis. Our data suggest the existence in several organs of the guinea pig of a widely distributed, yet discrete, system of venular endothelial cells specialized in the traffic of leukemic cells.     

Vascular invasion of the epithyseal growth plate: Analysis of metaphyseal capillary ultrastructure and growth dynamics

Metaphyseal blood vessels which invade the calcifying epiphyseal growth plate were examined by a variety of techniques to determine their morphology, cell division, and growth patterns as they relate to endochondral ossification. Four regions of these vessels were characterized: 1) sprout tips—the terminal ends of the capillary sprouts which impinge upon the hypertrophic chondrocytes of the growth plate; 2) region of extended calcified cartilage—those deeper vessels within the metaphysis which are surrounded by an extracellular matrix predominantly composed of extended septa of calcified cartilage; 3) region of bone deposition—further still from the epiphysis these microvessels are contained within a network of active bone deposition laid upon a scaffold of calcified cartilage; 4) region of primary vessels—at a distance of 350–500 μm from the epiphysis are dilated vessels with one or two layers of smooth muscle in their walls, which supply and drain the metaphyseal capillary plexus. The sprout tips are continuous blind-ended vessels lined with an attenuated endothelium with no underlying basement membrane. Dividing endothelial cells are most frequently found in the region of bone deposition 175–200 μm behind the apices of the growing sprout tips. A time-coursed, autoradiographic examination of cytokinesis revealed radio-labelled endothelial cells to appear at the epiphysis after a 24 hr period. The metaphyseal capillary sprouts represent a continuous, unidirectional angiogenic vascular network which grows by elongation from the region of bone deposition; this region remians a fixed distance behind the sprout tips. These findings are discussed in light of the growth dynamics between this vascular plexus and the epiphyseal growth plate.     

Morphologic observations on experimental corneal vascularization in the rat

The morphologic evolution of the early events of corneal vascularization in the rat cornea induced by silver nitrate cautery were followed by light and electron microscopy. An initial acute inflammatory response occurred within the first 6 hours after cautery as evidenced by vascular dilation, diapedesis of leukocytes, and an increased vascular permeability, as manifested by distended lymphatics and the presence of extravascular fibrin. At 24 hours after injury, the hypertrophy and the prominence of nucleoli and the presence of abundant polyribosomes were the initial suggestion that vascular endothelial cells and pericytes had entered the mitotic cycle. Nine hours later, at 33 hours after cautery, the first new vessels were observed as sprouts from the capillary arcade and postcapillary venules. The experiments reemphasize the association of acute inflammation and corneal vascularization in this model and underscore the early occurrence of vascular endothelial cell replication.     

Transmission and scanning-electronmicroscopy of experimental liver metastases derived from intrasplenically growing primary tumor

Liver metastasis formation from intrasplenically growing Lewis lung tumor was studied with transmission and scanning electron microscopy. Tumor cells arrested in the sinusoids formed desmosome-like junctions with endothelial cells and crossed the endothelial lining at intercellular gaps and by transcellular diapedesis. The metastatic foci had no newly formed vessels, and the stroma was provided by non-parenchymatous liver cells. Morphology suggested different outcome for tumor cell-host cell interactions at different stages of tumor growth. The host cell activity to destroy tumor cells was present only at the early stage but disappeared later, when the tumor cells were ready to phagocyte normal cells. The reason for this could be the immunosuppressive effect of the primary tumor. Results emphasize the importance to study metastasis formation in primary tumor-bearing hosts.     

Regional distributions of blood flow and tissue uptake rates of vitamin B12 and albumin within single rabbit skeletal muscles

The mechanism underlying the marked regional variations in blood flow within single skeletal muscles has not been identified. The present investigation was prompted by previous data pointing to a regional distribution of the number of perfused vessels as one possible determinant for the heterogeneous perfusion pattern. We examined this possibility by assessing the regional tissue uptake rates of vitamin B₁₂ and albumin assuming that these are related to the number of perfused vessels. This is the first study that has included simultaneous measurements of regional blood flow and of regional tissue uptake rates of vitamin B₁₂ and albumin within single skeletal muscles in the intact animal. The microsphere method was adopted to measure regional blood flow in 0.25 g muscle regions in awake and in anaesthetized rabbits. Various tracers were used to assess the regional distributions of tissue uptake rates and of vascular and interstitial volumes. Albeit displaying a marked regional heterogeneity, neither the tissue uptake rate of vitamin B₁₂ nor that of albumin were correlated to regional blood flow (P < 0.05) at rest (awake or anaesthetized rabbits) or during exercise hyperaemia (anaesthetized). At rest, but not during exercise, the distributions of regional tissue uptake rates of the two tracers showed a striking bimodal pattern. Vascular and interstitial volumes showed minimal interregional variations. The lack of correlation between regional blood flow and tissue uptake rates and the minute interregional variation in vascular volume, argue against variations in the number of perfused vessels as the explanation for the regional heterogeneity in blood flow.     

Structural and Functional Aspects of the Interaction of Inflammatory Cells with the Blood-Brain Barrier in Experimental Brain Inflammation

Interaction between various subclasses of inflammatory cells (ICs) and endothelial cells (ECs) lining selective blood vessels of the mammalian blood-brain barrier (BBB) is an initial, important event during inflammatory conditions of the central nervous system (CNS). In this review, we will present a brief ultracytochemical and immunocytochemical assessment of our perspective on this intimate cellular interaction which has been described recently in conditions that involve immunological alterations of the BBB. We will discuss some morphological aspects of what is currently known about acute and chronic inflammatory BBB disorders that are involved in inflammatory processes. We will focus, in particular, on experimental allergic encephalomyelitis (EAE), an animal model for human multiple sclerosis (MS). Many of the past and more recent concepts found in the literature concerning IC attachment and diapedesis, as well as our own experimental efforts over more than two decades will be presented.     

登革出血热皮疹的病理及超微结构研究

通过３例登革出血热皮疹处皮肤病理切片检查，对皮疹的发生、消退，病理改变，病变处血管等超微结构进行观察。结果显示登革出血热皮疹的主要病理改变为真皮上层血管内皮细胞的变性，致使红、白细胞外溢，推论内皮细胞的病变与病毒感染相关，并为出血的基本机理。     

In Vitro Expression of Adhesion Receptors and Diapedesis by Polymorphonuclear Neutrophils during Experimentally Induced Streptococcus uberis Mastitis

The expression of adhesion receptors and diapedesis by polymorphonuclear neutrophils (PMN) were studied before and during experimentally induced Streptococcus uberis mastitis. Both quarters of the left half of the udders of five midlactation cows were inoculated with a suspension containing approximately 500 CFU of S. uberis 0140J. Clinical signs of an inflammatory reaction and leukocyte influx were observed 24 h after challenge. The expression of CD11b/CD18 adhesion receptors, determined by flow cytometry, was upregulated 24 h after challenge. A confluent monolayer of bovine secretory mammary epithelial cells on collagen-coated inserts was used to study PMN diapedesis. Bovine C5a was used as the chemoattractant. An 80% decrease in PMN diapedesis was observed 24 h after challenge. The decrease in diapedesis continued for 3 weeks after challenge.     

Adhesion and signaling molecules controlling the transmigration of leukocytes through endothelium

Summary:  Migration of leukocytes into tissue is a key element of innate and adaptive immunity. While the capturing of leukocytes to the blood vessel wall is well understood, little is known about the mechanisms underlying the actual transmigration of leukocytes through the vessel wall (diapedesis). Even a basic question such as whether leukocytes migrate through openings between adjacent endothelial cells (junctional pathway) or through single endothelial cells (transcellular pathway) is still a matter of intensive debate. It is generally accepted that both pathways exist; however, whether they are of equal physiological significance is unclear. Several endothelial adhesion and signaling molecules have been identified, most of them at endothelial cell contacts, which participate in leukocyte diapedesis. A concept is evolving suggesting that transendothelial migration of leukocytes is a stepwise process. Blocking or eliminating some of the different adhesion and signaling proteins results in very different effects, such as trapping of leukocytes above endothelial cell contacts, in between endothelial cells, or between the endothelium and the underlying basement membrane. Other proteins are involved in the opening of endothelial cell contacts and yet others in their maintenance providing the barrier for extravasating leukocytes. The various molecular players and the functional steps involved in diapedesis are discussed.     

Leukocyte-endothelial-cell interactions in leukocyte transmigration and the inflammatory response

Both the innate and adaptive immune responses are dependent on the migration of leukocytes across endothelial cells. The process of diapedesis, in which the leukocyte crawls between tightly apposed endothelial cells, is a unique and complex process. Several molecules concentrated at the junctions of endothelial cells, originally described as having a role in holding the endothelial monolayer together, have also been shown to have a role in the emigration of leukocytes. Several mechanisms have been proposed for 'loosening' the junctions between endothelial cells to enable leukocyte passage. These leukocyte-endothelial-cell adhesion molecules are probably involved in regulating the signaling as well as the adhesion events of diapedesis. In addition, this Review introduces a new and unified nomenclature for the junctional adhesion molecule (JAM) family.     

Lymphatic capillaries of the pig lung: TEM and SEM observations

Pulmonary lymphatic vessels extend within the connective tissue sheets surrounding airways and blood vessels. Frequently in this location they also border the lobular parenchyma, but no lymphatic vessels have been found within intralobular compartments between blood capillaries and alveoli. The presence and distribution of lymphatic vessels in pulmonary tissue are consistent with an important role for the lymphatic system in the clearance of interstitial fluids in the lung. Pulmonary lymphatic channels have structural characteristics of initial lymphatics; their walls are formed only by an endothelial layer, and no muscular cells are present. A network of anchoring filaments and collagen and elastic fibers surrounds the vessel walls. Because the lung is a mobile organ the tissue undergoes compression and distension during respiratory phase. These modifications could have a role in the mechanisms for lymph formation and flow. © 1994 Wiley-Liss, Inc.     

Ethanol and tolerated doses ofAmanita phalloides protect against lethal doses of the mushroom

Single tolerated doses of a lyophilizate fromAmanita phalloides (APL) failed to increase the survival rate of mice after the LD₉₀ of the mushroom. Also, pretreatment with large doses of ethanol did not protect against lethal doses of APL. However, with combined administration of single tolerated doses of APL and of ethanol, the survival rate of mice after a lethal challenge with APL was increased from 13% to up to 100%. Both ethanol and APL potentiated the effect of barbiturate narcosis, but the combined pretreatment was not more effective. The protection against APL afforded by the combined regimen may be due to inhibition of activating hepatic microsomal enzymes, but biochemical studies are needed to support this hypothesis.     

Isolation of the structural polypeptides of foot-and-mouth disease virus and analysis of their C-terminal sequences

The three polypeptides—VP₁, VP₂ and VP₃—which comprise approximately 91% of the protein of foot-and-mouth disease virus, type A₁₂-119, were separated by polyacrylamide gel electrophoresis (PAGE) in sufficient purity and quantity for determination of their amino acid compositions as well as for analysis of their C-terminal sequences and molecular weights by reaction with carboxypeptidase-A. At least seven distinct differences in amino acid compositions were observed among the three polypeptides, but their average composition was nearly identical to that of the total protein of the virus, VP_0–4. C-Terminal amino acid sequences for VP₁ and VP₃ appear to to be (—serine-glutamine) and (—glutamine-alanine-leucine), respectively, and the C-terminal amino acid of VP₂ is probably glutamic acid. Molecular weights for VP₁ and VP₃ calculated from C-terminal data were in the 27,500–31,000 dalton range compared with values of 28,500 determined previously by PAGE.     

Leukocyte podosomes sense their way through the endothelium

Whether leukocytes can transmigrate through endothelial cells, rather than between them, is controversial. In this issue, Carman et al. (2007) demonstrate that lymphocytes can extend podosomes to palpate endothelial cells searching for areas permissive for transcellular diapedesis.