[Microcirculation disorders in shock].
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Humans↗
Biomedical subjects
K Ley
Publications and source records attributed to K Ley.
A versatile intravital microscope design.
A new intravital microscope system with a versatile arrangement of optical elements and different photosensitive detection devices is described. The image formed by the microscope modified for telescopic imaging is transferred to an optical platform. Through an arrangement of beam splitters and mirrors, the image can be projected simultaneously into three of four different recording systems, including video- and photocameras and a velocity measuring system. In contrast to earlier approaches, the presented system simultaneously generates images on several devices. These images have the same orientation and field of view as that seen through the oculars. This allows the parallel application of different measuring techniques to a given area of the microcirculation.
Humans↗
Segmental differences of microvascular permeability for FITC-dextrans measured in the hamster cheek pouch.
An intravital microscopic method for quantitative measurement of interstitial concentrations of fluorescent tracers has been applied to the investigation of microvascular permeability in the hamster cheek pouch. Some nanoliters of FITC-dextran mean mol wt (Mw) 20,000, 3000, or sodium fluorescein (Mw 376) were injected into an arteriole of the exposed cheek pouch via micropipet. The extravasation of fluorochromes was measured by a photodensitometric method including two sets of calibration procedures (in vitro and in vivo). Postcapillary and collecting venules exhibited the highest absolute increase of fluorochrome concentration in the tissue for all tracer molecules tested when compared to arterioles or capillaries. The permeability of the vascular wall was quantified, assuming that diffusion processes play the main role for the transport of the investigated molecules under the experimental conditions of a high concentration gradient across the membrane. Permeability coefficients P (cm/sec) and apparent diffusion coefficients D' (cm2/sec) of the microvascular wall were calculated using a mathematical model for one-dimensional diffusion in composite media. The analysis is based on measured data of interstitial diffusion coefficients of the tracers used. For all tracer molecules tested, the wall of the capillaries and postcapillary venules was significantly more permeable than the arteriolar wall. For the largest test molecule (FITC-dextran Mw 20,000), the permeability coefficient of the vessel wall showed a maximum in the postcapillary venules. These findings support the concept of a "gradient of permeability" with a nonuniform distribution of exchange capacity only for the precapillary microvessels. A marked preponderance of venular over capillary permeability could, if at all, only be detected for FITC-dextran Mw 20,000. The present study characterizes the vessel wall by apparent diffusion coefficients which are, for FITC-dextran Mw 3000, and free fluorescein, roughly three orders of magnitude lower than the apparent diffusion coefficients in connective tissue.
Animals↗
Topological structure of rat mesenteric microvessel networks.
Microvascular lengths, diameters, and flow directions were determined in all vessel segments (n = 1303) between bifurcations in three complete rat mesenteric microvessel networks (25 mm2 each) using intravital video- and photomicroscopy. The classification of vessel segments as arteriolar, venular, or av-segments (all segments connecting the arteriolar to the venular tree) was based on purely topological criteria. The topological structure of the networks was analyzed using the Horton-Strahler technique and a new generation scheme. Generation numbers were assigned to the vessel segments on the basis of the number of upstream (in the arteriolar tree) and downstream (in the venular tree) bifurcations. The mean generation number of the av-segments, a characteristic parameter of the generation scheme, reflects the topological structure of the network more accurately than Horton's branching ratio Rb. Both the arteriolar and venular tree of the mesenteric networks were found to be dichotomous branching structures which were neither strictly symmetric nor strictly asymmetric. The topological information obtained was compared to network models generated by different random branching algorithms. The result of this comparison suggests that the network structure changes at a certain generation level. Distal to this generation level, the mesenteric networks resemble a model network generated by random branching at any segment, while the proximal portion is similar to a model allowing random branching at terminal segments only.
Animals↗
Generalization of the Fahraeus principle for microvessel networks.
Microvessel hematocrits and diameters were determined in each vessel segment between bifurcations of three complete microvascular networks in rat mesentery. Classification of the segments as arteriolar, venular, or arteriovenular (av) was based on flow direction at branch points. Photographic and videomicroscopic mapping was used to obtain quantitative information on the architecture and topology of the networks. This topological information allowed the analysis of hematocrit distribution within a series of consecutive-flow cross sections, each of which carried the total flow through the network. The observed reduction of mean hematocrit in the more peripheral cross sections is explained by the presence of a "vessel" and a "network" Fahraeus effect. The vessel Fahraeus effect results from velocity difference between red cells and blood within the individual vessel segments due to the existing velocity and cell concentration profiles. The network Fahraeus effect is based on the velocity difference of red cells and blood caused by velocity and hematocrit heterogeneity between the vessels constituting any of the complete-flow cross sections. The network Fahraeus effect is found to account for approximately 20% of the total hematocrit reduction and increases toward the most distal cross sections.
Animals↗
Microvascular permeability and blood flow in atrial homografts in the hamster cheek pouch.
Atrial heart tissue grafts from newborn golden hamsters were transplanted into cheek pouches and were studied using intravital microscopic techniques. The smallest microvessels in the homografts showed a mean diameter of 13.3 +/- 1.1 micron (mean +/- S.E.M.) thus being in the dimensional range of terminal arterioles of the fourth branching order (A4, 14.0 +/- 2.1 micron) in normal cheek pouch tissue. Arteriolar red cell velocity was measured using a modified dual slit method. Volumetric blood flow was calculated and seen to be similar in terminal cheek pouch arterioles (116 +/- 34 pl/s) and in the graft vessels (119 +/- 26 pl/s). The responsiveness to topically applied norepinephrine of the graft vessels was found to be significantly lower than in cheek pouch arterioles. Microvascular permeability studied with FITC-dextrans (intravenous application) revealed significantly higher intensity values in the homografts as compared with the surrounding cheek pouch tissue. The difference was most pronounced for the largest tracer molecules used (Mw 150 000). This phenomenon is most probably due to an increased macromolecular permeability of the microvessels in the graft. The present findings point towards functional differences between the transplant vessels and cheek pouch capillaries.
Animals↗
Morphological and functional aspects of heart homograft adaptation to hamster cheek pouch. Spontaneous pulsatile activity and ultrastructure.
Atrial homografts from newborn hamsters, transplanted into cheek pouches of adult Syrian golden hamsters, were studied using intravital and electron microscopy. Half of the transplants survived up to 1 year, performing spontaneous pulsatile activity. The transplants could be influenced by beta-receptor activating or blocking substances applied intravenously to the host animal. For morphological analysis homografts were taken 3, 6, and 12 months after transplantation. Irregular arrangement of the myofibers occurred, and the T system was seen to be replaced by plasmalemmal and sub-plasmalemmal vesiculations. There was some evidence for hypoxia in the grafts. Dedifferentiation of atrial cardiocytes, somehow resembling smooth muscle cells, is discussed. Despite these morphological changes the specific contractile function of the grafts was preserved.
Animals↗
Changes in macromolecular permeability by intravascular generation of oxygen-derived free radicals.
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Animals↗
Migrant women: is migration a blessing or a handicap? Situation of migrant women in Switzerland.
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Demography↗
Targeted ultrasound contrast agent for molecular imaging of inflammation in high-shear flow.
Targeted ultrasound contrast materials (gas-filled microbubbles carrying ligands to endothelial selectins or integrins) have been investigated as potential molecular imaging agents. Such microbubbles normally exhibit good targeting capability at the slower flow conditions. However, in the conditions of vigorous flow, binding may be limited. Here, we describe a microbubble capable of efficient binding to targets both in slow and fast flow (exceeding 4 dyne/cm(2) wall shear stress) using a clustered polymeric form of the fast-binding selectin ligand sialyl Lewis(X). Microbubbles were prepared from decafluorobutane gas and stabilized with a monolayer of phosphatidylcholine, PEG stearate and biotin-PEG-lipid. Biotinylated PSLe(x) (sialyl Lewis(X) polyacrylamide) or biotinylated anti-P-selectin antibody (RB40.34) was attached to microbubbles via a streptavidin bridge. In a parallel plate flow chamber targeted adhesion model, PSLe(x) bubbles demonstrated specific adhesion, retention and slow rolling on P-selectin-coated plates. Efficiency of firm targeted adhesion to a P-selectin surface (140 molecules/microm(2)) was comparable for antibody-carrying bubbles and PSLe(x)-targeted bubbles at 0.68 dyne/cm(2) shear stress. At fast flow (4.45 dyne/cm(2)), PSLe(x)-targeted bubbles maintained their ability to bind, while antibody-mediated targeting dropped more than 20-fold. At lower surface density of P-selectin (7 molecules/microm(2)), targeting via PSLe(x) was more efficient than via antibody under all the flow conditions tested. Negative control casein-coated plates did not retain bubbles in the range of flow conditions studied. To confirm echogenicity, targeted PSLe(x)-bubbles were visualized on P-selectin-coated polystyrene plates by ultrasound imaging with a clinical scanner operated in pulse inversion mode; control plates lacking targeted bubbles did not show significant acoustic backscatter. In vivo, in a murine model of inflammation in the femoral vein setting, targeting efficacy of intravenously administered PSLe(x)-microbubbles was comparable with targeting mediated by anti-P-selectin antibody, and significantly exceeded the accumulation of non-targeted control bubbles. In the inflamed femoral artery setting, PSLe(x)-mediated microbubble targeting was superior to antibody-mediated targeting.
Acrylic Resins↗
Selectin ligands promote ultrasound contrast agent adhesion under shear flow.
Contrast-enhanced ultrasound imaging has shown promise in the field of molecular imaging. This technique relies upon the adhesion of ultrasound contrast agent (UCA) to targeted molecular markers of disease. This is accomplished by coating the surface of the contrast agent with a ligand that specifically binds to the intended molecular marker. Most UCA particles remain in the blood space, and their retention is influenced by the forces imposed by blood flow. For a UCA bound to a molecular target on the vascular endothelium, blood flow imposes a dislodging force that counteracts retention. Additionally, contrast agent adhesion to the molecular marker requires rapid binding kinetics, especially in rapid blood flow. The ability of a ligand:target bond complex to mediate fast adhesion and withstand dislodging force is necessary for efficient ultrasound-based molecular imaging. In the current study, we describe a flow-based adhesion assay which, combined with a novel automated tracking algorithm, enables quick determination of the ability of a targeting ligand to mediate effective contrast agent adhesion. This system was used to explore the adhesion of UCA targeted to the proinflammatory endothelial protein P-selectin via four targeting ligands, which revealed several interesting adhesive behaviors. Contrast agents targeted with glycoconjugate ligands modeled on P-selectin glycoprotein ligand 1 exhibited primarily unstable or transient adhesion, while UCA targeted with an anti-P-selectin monoclonal antibody exhibited primarily firm adhesion, although the efficiency with which these agents were recruited to the target surface was relatively low.
Adhesiveness↗
L-selectin: mechanisms and physiological significance of ectodomain cleavage.
L-selectin is a cell adhesion molecule consisting of a large, highly glycosylated, extracellular domain, a single spanning transmembrane domain and a small cytoplasmic tail. It is expressed on most leukocytes and is involved in their rolling on inflamed vascular endothelium prior to firm adhesion and transmigration. It is also required for the constitutive trafficking of lymphocytes through secondary lymphoid organs. Like most adhesion molecules, L-selectin function is regulated by a variety of mechanisms including gene transcription, post-translational modifications, association with the actin cytoskeleton, and topographic distribution. In addition, it is rapidly downregulated by proteolytic cleavage near the cell surface by ADAM-17 (TACE) and at least one other "sheddase". This process of "ectodomain shedding" results in the release of most of the extracellular portion of L-selectin from the cell surface while retaining the cytoplasmic, transmembrane, and eleven amino acids of the extracellular domain on the cell. This review will examine the mechanism(s) of L-selectin ectodomain shedding and discuss the physiological implications.
ADAM Proteins↗
Arterial injury increases expression of inflammatory adhesion molecules in the carotid arteries of apolipoprotein-E-deficient mice.
Recent studies demonstrate increased cellular adhesion molecule expression by neointimal endothelium overlying primary and restenotic atherosclerotic plaque. In this study, we developed an atherosclerotic mouse model of arterial injury and characterized adhesion molecule expression after injury. Sixteen apolipoprotein-E-(ApoE)-deficient mice fed a Western-type diet for 4 weeks underwent carotid artery wire denudation at week 2. For each segment, the extent of neointima formation and medial thickening, or adhesion molecule expression, were scored separately on a scale from 0 (no plaque/thickening or expression) to 3 (extensive plaque/thickening or expression) using Movat staining (n = 3) or immunohistochemical analysis (n = 13). Histology revealed significant medial thickening (1.8 +/- 0.9 vs. 0.3 +/- 0.5, p < 0. 001) versus controls and pronounced staining for monocytes/macrophages in the wall of injured vessels. Immunohistochemical analysis showed more robust expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) on the luminal surface of injured arteries versus controls (2.2 +/- 0.6 vs. 1.4 +/- 0.7, p < 0.01, and 2.5 +/- 0.5 vs. 1.2 +/- 0.6, p < 0.001, respectively). Injury increased adventitial ICAM-1 expression (2.6 +/- 0.5 vs. 1.6 +/- 0.5, p < 0.002) and medial VCAM-1 expression (2.2 +/- 0.6 vs. 1.2 +/- 0. 7, p < 0.004). Thus, carotid injury results in significant medial thickening and increases adhesion molecule expression beyond that induced in ApoE-deficient mice fed a Western diet alone. The observation of macrophage infiltration into the media at sites of increased ICAM-1 and VCAM-1 expression suggests that these molecules may mediate monocyte/macrophage trafficking into the wall of injured arteries.
Animals↗
Role of primary and secondary capture for leukocyte accumulation in vivo.
Leukocyte accumulation during inflammation depends on the concerted action of selectin and integrin adhesion molecules, which promote capture, rolling, and arrest of these cells on activated endothelium. In addition to interacting with endothelial cells, leukocytes can also adhere to already adherent leukocytes through an L-selectin-dependent mechanism. Initiation of adhesion through this mechanism has been called nucleation and leads to characteristic geometric patterns (ie, clusters and strings) of adherent leukocytes in flow chambers. We have used intravital microscopy of tumor necrosis factor-alpha (TNF-alpha)-treated mouse cremaster muscles to quantitatively investigate the potential role of leukocyte-leukocyte adhesion in initiating and maintaining the leukocyte clusters that are commonly observed in inflamed venules. Our data show that in TNF-alpha-treated venules with diameters between 23 and 108 microm, leukocyte adhesion occurs in clusters that are 19 to 50 microm long and 8 to 44 microm wide. They are almost entirely made up of slow-rolling leukocytes. Of all leukocytes recruited into a cluster (100%), the majority enter the cluster rolling along the endothelium and sharply reduce their velocity in the absence (59%) or presence (15%) of other leukocytes in proximity (one cell diameter). Some of the rolling leukocytes (17%) pass through the cluster without reducing their velocity. Recruitment of leukocytes from the free flow regime into a cluster is a rare event and accounts for only 7 (1.2%) of 476 leukocytes arriving in the cluster. However, of the leukocytes captured from the free flow, 6 initiated contact with a slow-rolling leukocyte rather than making direct contact with the endothelium. Our data show that leukocyte-leukocyte interactions can occur in vivo but are not important for cluster formation. This is confirmed by the observation of normal cluster formation in L-selectin-deficient mice, in which leukocyte-leukocyte interactions under flow are abolished. We conclude that leukocyte-mediated nucleation contributes little to leukocyte recruitment during inflammation in vivo. Cluster formation appears to be dominated by areas of endothelium with a higher expression of E-selectin, because cluster formation is greatly reduced in E-selectin-deficient mice.
Animals↗
The New York State Experiment.
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Adaptation, Physiological↗