Biomedical subjects

Ingrid H Sarelius

Publications and source records attributed to Ingrid H Sarelius.

14 recordsLinked to original sources

PubMed2006

TNF-alpha activation of arterioles and venules alters distribution and levels of ICAM-1 and affects leukocyte-endothelial cell interactions.

The observation that leukocyte-endothelial cell (EC) interactions are localized to specific regions on the microvessel wall suggests that adhesion molecule distribution is not uniform. We investigated ICAM-1 distribution and leukocyte-EC interactions in blood-perfused microvessels (<80 mum) in cremaster muscle of anesthetized mice, using intravital confocal microscopy and immunofluorescent labeling. Variability of ICAM-1 expression directly determines leukocyte adhesion distribution within the venular microcirculation and contributes to leukocyte rolling in arterioles during inflammation. The number of rolling interactions increased with ICAM-1 intensity (r(2) = 0.69, P < 0.05), and rolling velocity was lower in regions of higher ICAM-1 intensity. In controls, venular ICAM-1 expression was approximately twofold higher than in arterioles. After TNF-alpha treatment, ICAM-1 expression was significantly increased, 2.8 +/- 0.2-fold in arterioles and 1.7 +/- 0.2-fold in venules (P < 0.05). ICAM-1 expression on activated arteriolar ECs only reached the level of control venular ICAM-1. Arteriolar but not venular ECs underwent redistribution of ICAM-1 among cells; some cells increased and some decreased ICAM-1 expression, magnifying the variability of ICAM-1. TNF-alpha treatment increased the length of bright fluorescent regions per unit vessel length (42%, control; 70%, TNF-alpha) along the arteriolar wall, whereas no significant change was observed in venules (60%, control; 63%, TNF-alpha). The spatial distribution and expression levels of adhesion molecules in the microcirculation determine the timing and placement of leukocyte interactions and hence significantly impact the inflammatory response. That arteriolar ECs respond to TNF-alpha by upregulation of ICAM-1, although in a different way compared with venules, suggests an explicit role for arterioles in inflammatory responses.

Animals↗

PubMed2005

Macromolecule permeability of in situ and excised rodent skeletal muscle arterioles and venules.

In microvessels, acute inflammation is typified by an increase in leukocyte-endothelial cell interactions, culminating in leukocyte transmigration into the tissue, and increased permeability to water and solutes, resulting in tissue edema. The goal of this study was to establish a method to quantify solute permeability (P(s)) changes in microvessels in intact predominantly blood-perfused networks in which leukocyte transmigratory behavior could be precisely described using established paradigms. We used intravital confocal microscopy to measure solute (BSA) flux across microvessel walls, hence P(s). A quantitative fluorescence approach (Huxley VH, Curry FE, and Adamson RH. Am J Physiol Heart Circ Physiol 252: H188-H197, 1987) was adapted to the imaged confocal tissue slice in which the fluorescent source volume and source surface area of the microvessel were restricted to the region of vessel that was contained within the imaged confocal tissue section. P(s) measurements were made in intact cremaster muscle microvasculature of anesthetized mice and compared with measurements of P(s) made in isolated rat skeletal muscle microvessels. Mouse arteriolar P(s) was 9.9 +/- 1.1 x 10(-7) cm/s (n = 16), which was not different from 8.4 +/- 1.3 x 10(-7) cm/s (n = 6) in rat arterioles. Values in venules were significantly (P < 0.05) higher: 44.4 +/- 7.9 x 10(-7) cm/s (n = 14) in mice and 25.0 +/- 3.7 x 10(-7) cm/s in rats. Convective coupling was estimated to contribute <10% to the measured P(s) in both microvessel types and both animal models. We conclude that this approach provides an appropriate quantification of P(s) in the intact microvasculature and that arteriolar P(s), while lower than in venules, is nevertheless consistent with arterioles being a significant source of interstitial protein.

Animals↗

PubMed2005

Preferential binding of leukocytes to the endothelial junction region in venules in situ.

OBJECTIVE: To determine whether leukocytes show a preference to firmly adhere to the endothelium near the endothelial cell (EC) junction in their native environment, i.e., blood-perfused venules in situ. METHODS: Intravital confocal microscopy was used to determine the positions of firmly adherent leukocytes with respect to EC junctions in cremaster muscle venules in anesthetized mice. EC area and EC junction locations were identified using immunofluorescent labeling of platelet-endothelial cell adhesion molecule-1 (PECAM-1), an endothelial junction protein. Leukocytes were identified using transillumination through the same optical path. RESULTS: Seventy-five percent of firmly adherent leukocytes overlapped an EC junction (the average distance to the nearest EC junction was 2.4 +/- 0.1 micro m, n = 263). This distance was less in smaller diameter venules, hence the percentage of adherent leukocytes that overlapped an endothelial junction was larger. EC shape and size varied with venular diameter: in smaller venules (30-49 micro m diameter), ECs had significantly less area (316 +/- 19 micro m(2)) and were narrower (12.5 +/- 0.4 micro m) than in larger (70-89 micro m diameter) venules, which had 614 +/- 28 micro m(2) area and 17.8 +/- 0.5 micro m width, respectively (p < .001). Furthermore, the majority (73.0%) of firmly adherent leukocytes crawled an average distance of 29.4 +/- 2.8 micro m at a mean intralumenal migration velocity of 7.6 +/- 0.4 micro m/min before undergoing transmigration or detaching and reentering the free fluid stream. CONCLUSIONS: Most adherent leukocytes in blood-perfused venules are located at or near EC junctions. This is primarily due to the size and shape of venular ECs. Further, most leukocytes that appear to be adherent are in fact motile, and migrate a significant distance along the lumenal wall. The authors speculate that this reflects movement to those EC junctional regions that support transendothelial migration.

Animals↗

PubMed2005

The distribution of rolling neutrophils in venular convergences.

The aim of this study was to characterize the distribution of adherent leukocytes in branched venular convergences in vivo. Intravital microscopy was used to obtain video images of leukocyte adhesion in multiple branched sites in mouse cremaster muscle, during the mild inflammatory response induced by surgical preparation. The average number of cells/vessel length was obtained over several minutes for seven venular convergences with varying geometrical configurations. Results from this study demonstrate a strong tendency of leukocytes to adhere at junctional points between converging vessels. Different vessel configurations were studied and results were shown to be insensitive to precise vessel geometry. Thus, in post-capillary venules, leukocytes are most likely to adhere at points between converging vessels, regardless of the precise geometrical properties or configuration of the vessels. Hydrodynamic mechanisms due to flow behavior through convergences likely play a significant role in determining locations of cellular adhesion. Future work should concentrate on quantifying the relative contributions of hydrodynamic and biochemical mechanisms to aid in understanding disease processes and development of treatments or therapeutics.

Animals↗

PubMed2004

Role of shear forces and adhesion molecule distribution on P-selectin-mediated leukocyte rolling in postcapillary venules.

Rolling on the venular endothelium is a critical step in the recruitment of leukocytes during the inflammatory response. P-selectin is a key mediator of leukocyte rolling, which is an early event in the inflammatory cascade; this rolling is likely to be directly regulated by both local fluid shear forces and P-selectin site densities in the microvasculature. However, neither the spatial pattern of P-selectin expression in postcapillary venules nor the effect of local expression patterns on rolling behavior in intact functional venules is known. We investigated the influence of local shear forces and the spatial distribution of endothelial P-selectin in intact blood perfused post capillary venules in anesthetized mice using intravital confocal microscopy, high temporal resolution particle tracking, and immunofluorescent labeling. We demonstrated a shear-dependent increase in average leukocyte rolling velocity that was attributable to a shear-dependent increase in the occurrence of transient leukocyte detachments from the endothelial surface: translational velocity during leukocyte contact with the vessel wall remained constant. P-selectin expression was not different in venules with characteristically different shear rates or diameters but varied significantly within individual venules. In postcapillary venules, regions of high P-selectin expression correlated with regions of slow leukocyte rolling. Thus the characteristically variable leukocyte rolling in vivo is a function of the spatial heterogeneity in P-selectin expression. The study shows how the local hydrodynamic forces and the nonuniform pattern of P-selectin expression affect the behavior of interacting leukocytes, providing direct evidence for the local variation of adhesion molecule expression as a mechanism for the regulation of leukocyte recruitment.

Animals↗

PubMed2004

Localized transient increases in endothelial cell Ca2+ in arterioles in situ: implications for coordination of vascular function.

Intracellular Ca2+ transients were identified in endothelial cells (ECs) in intact blood-perfused arterioles. ECs in cremaster muscle arterioles (diameter approximately 45 microm) in anesthetized mice were loaded with the Ca2+ indicator fluo 4-AM by intraluminal perfusion, after which blood flow was reestablished. Confocal microscopy was used to visualize Ca2+ as a function of fluo-4 intensity in real time. Separate sets of experiments were performed under the following conditions: control, ischemia, during inhibition of P(2x) or P(1) purinoreceptors, and with the application of exogenous adenosine. In controls, spontaneous EC Ca2+ transients displayed a wide range of activity frequency (1-32 events/min) and about one-third of these transient events were synchronized between adjacent ECs. The increase in Ca2+ remained localized and did not spread to encompass the entire cell body. Ca2+ transient activity decreased significantly with ischemia (from 9.9 +/- 0.6 to 3.1 +/- 0.3 events/min, n = 135) but was unaffected by P(2x) or P(1) receptor inhibition. Exogenous adenosine significantly increased the frequency of Ca2+ transients (to 12.8 +/- 0.9 events/min) and increased synchronization so that 50% of all Ca2+ events were synchronized between ECs. This response to adenosine was not due to an increase in shear stress. These data indicate that localized Ca2+ transients are sensitive to flow conditions and, separately, to metabolically active pathways (exogenous adenosine), although the basal activity occurs independently of P(2x) or P(1) receptors. These transients may represent a mechanism by which individual EC responses are integrated to result in coordinated arteriolar responses in situ.

Adenosine↗

PubMed2004

The effect of hematocrit and leukocyte adherence on flow direction in the microcirculation.

We sought to characterize how adherent leukocytes at the vessel wall, and the presence of erythrocytes, alter the streamlines (paths) of blood flow in the postcapillary venules. We directly visualized blood flow and leukocyte-endothelial cell interactions in postcapillary venules located in the cremaster muscle of anesthetized mice. Fluid streamlines were visualized by perfusing the cremaster muscle tissue with 0.5-micron fluorescent beads suspended in either buffer or whole blood, to examine the effect that erythrocytes have on the directionality of flow. Acute inflammation was induced in some animals by pretreatment of the vessels with tumor necrosis factor-alpha. To quantify the flow direction, the average deflection angle was defined as a scalar metric. Tracer bead trajectories were measurably altered by the presence of systemic levels of hematocrit, determined in each animal to be about 45%. Deviation from undirectional flow was also found to: (i) decrease with increasing vessel diameter, and (ii) increase with the number of adherent leukocytes. Fluid streamlines in the presence or absence of leukocyte adhesion or red cells agreed qualitatively with those obtained from theoretical calculations of blood flow using multiparticle adhesive dynamics. The microscale characteristics of venular flow are significantly altered during inflammation or changes in local hematocrit.

Animals↗

PubMed2003

Increase in endothelial cell Ca(2+) in response to mouse cremaster muscle contraction.

We addressed the role of endothelial cells (ECs) in metabolic dilatation of skeletal muscle arterioles in anaesthetized mice in situ. Electrical field stimulation was used to contract the cremaster muscle for 15 s at 30 Hz. Diameter was observed using bright field microscopy. In controls, muscle contraction produced a 15.7 +/- 1.5 microm dilatation from a baseline of 17.4 +/- 1.6 microm. Endothelial denudation (-EC) via intraluminal perfusion of air abolished this response (1.6 +/- 1.2 microm in -EC, P < 0.05), identifying endothelium as the primary vascular cell type initiating the dilatation. To investigate the role of EC Ca(2+) in metabolic dilatation, arteriolar ECs were loaded with Fluo-4 AM or BAPTA AM by intraluminal perfusion, after which blood flow was re-established. Ca(2+) activity of individual ECs was monitored as a function of change from baseline fluorescence using confocal microscopy. In ECs, whole cell Ca(2+) increased (>10%, P < 0.05) during muscle contraction, and localized Ca(2+) transients were increased (>20%, P < 0.05) during the first minute after contraction. Chelation of EC Ca(2+) abolished the dilatations in response to muscle contraction (1.1 +/- 0.7 microm, P < 0.05). Inhibition of P(1) purinergic receptors (with xanthine amine congener) did not alter the rate of onset of the dilatation (P > 0.05) but decreased its magnitude immediately post stimulation (7.1 +/- 0.9 microm, P < 0.05) and during recovery. These findings demonstrate obligatory roles for endothelium and EC Ca(2+) during metabolic dilatation in intact arterioles. Furthermore, they suggest that at least two separate pathways mediate the local response, one of which involves stimulation of endothelial P(1) purinergic receptors via endogenous adenosine produced during muscle activity.

Adenosine↗

PubMed2003

Conducted dilations initiated by purines in arterioles are endothelium dependent and require endothelial Ca2+.

The signaling pathways underlying the regulation of vascular resistance by purines in intact microvessels and particularly in communication of remote vasomotor responses are unclear. One process by which remote regions of arterioles communicate is via transmission of signals axially along the vessel wall. In this study, we identified a pathway for local and conducted dilations initiated by purines. Adenosine (Ado) or ATP (bind P1 and P2 purinergic receptors, respectively) was micropipette applied to arterioles (maximum diameter approximately 40 microm) in the cheek pouch of anesthetized hamsters. Observations were made at the site of stimulation (local) or approximately 1200 microm upstream along the same vessel. P2 antagonists (pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid tetrasodium and suramin) inhibited local constriction to ATP, whereas local and upstream dilations were unaffected. In contrast, during inhibition of P1 receptors (with xanthine amine congener) the local constriction was unchanged, whereas both local and upstream dilations to ATP were inhibited. Hydrolysis of ATP to Ado is implicated in the dilator response as blocking 5'-ectonucleotidase (with alpha,beta-methyleneadenosine 5'-diphosphate) attenuated ATP-induced dilations. After endothelium denudation, constriction to ATP was unchanged, but dilations to both ATP and Ado were inhibited, identifying endothelial cells (ECs) as the primary target for P1-mediated dilation. Purines increased EC Ca2+ locally and upstream. Chelation of EC Ca2+ (with BAPTA) abolished the local and upstream dilations to P1 receptor stimulation. Collectively, these data demonstrate that stimulation of P1 receptors on ECs produces a vasodilation that spreads to remote regions. There is an associated increase in EC Ca2+, which is a required signaling intermediate in the manifestation of both the local and axially communicated arteriolar dilations.

5'-Nucleotidase↗

PubMed2003

Distributions of wall shear stress in venular convergences of mouse cremaster muscle.

OBJECTIVE: Wall shear stress regulates a variety of vascular functions and can be affected by variations in blood viscosity and wall shear rate independently. Therefore, the distribution of wall shear stress was characterized in converging flow regions of postcapillary venules to identify the relative contributions of shear rate and viscosity to wall shear rate and to determine whether venular branching directly affects the local shear environment. METHODS: Blood viscosity was evaluated as a function of vessel diameter and hematocrit, determined from fluorescent red blood cell (RBC) flux. Wall shear rate was measured directly from velocity profiles acquired using fluorescent RBC tracking or was estimated assuming Poiseuille flow conditions. Wall shear stress was calculated for measurements made across regions of converging flow. RESULTS: Measurements of wall shear stress showed no significant variation with location across the convergence (p < 0.9). Measures of viscosity showed little variation, for local measures of hematocrit (2.9 +/- 0.1 cp; coefficient of variation [CV]: 12%) or systemic hematocrit (3.4 +/- 0.1 centipoise [cp]; CV: 14%). Measures of wall shear rate showed greater variability for both direct measurements (61.2 +/- 8.7s; CV: 92%) and estimates assuming Poiseuille flow (122.7 +/- 15.7 s; CV: 83%). CONCLUSIONS: Variations in wall shear stress are independent of location through a venular convergence in contrast to arteriolar bifurcations. In addition, wall shear rate is significantly more variable than viscosity and, therefore, primarily accountable for wall shear stress variations in postcapillary venules.

Animals↗

PubMed2003

Hydrodynamic interactions between rolling leukocytes in vivo.

OBJECTIVE: The aim of this study was to characterize the hydrodynamic interactions between rolling and free-stream leukocytes in an in vivo model of selectin-mediated rolling, and to identify those physical mechanisms that influence the dynamics of transient adhesion with the walls of postcapillary venules. METHODS: Postcapillary venules of diameter 22-37 microm in the cheek pouch of anesthetized hamsters were visualized using intravital microscopy, with selectin-mediated rolling occurring in response to surgical preparation. RESULTS: Rolling velocity was found to be a strong function of the center-to-center separation with the nearest cell, and also was found to correlate strongly with the number of nearby cells. These effects are shown to be beyond that attributable to variations along the length of the vessel. Adherent leukocytes were observed to provide a nucleation site, precipitating further adhesion events of free-stream cells. CONCLUSIONS: The dynamics of the transient adhesion of leukocytes to the vessel wall in postcapillary venules is strongly dependent on the local concentration of adherent leukocytes, due to the complex hydrodynamics induced by their presence. The results are shown to agree well with theoretical considerations of the flow field induced by multiple nearby cells, suggesting a need for the future exploration of multicellular effects in the microcirculation.

Animals↗

PubMed2002

Muscle contraction under capillaries in hamster muscle induces arteriolar dilatation via K(ATP) channels and nitric oxide.

We tested the hypothesis that adenosine and nitric oxide can be sensed by capillaries and are implicated in the remote arteriolar dilatation initiated by muscle contraction. We also explored a role for K(ATP) channel activity in this response. Small bundles of muscle fibres underlying a group of capillaries in cremaster muscles of anaesthetized hamsters were electrically stimulated to contract for 2 min at each of 2, 4 and 8 Hz. Diameter changes were measured in the inflow arteriole to the group of capillaries after muscle contraction in the presence or absence of 10(-6) M xanthine amine congener (XAC) to block A(1) and A(2) adenosine receptors, 10(-4) or 10(-3) M N(omega)-nitro-L-arginine (LNNA) to block nitric oxide production, or 10(-5) M glibenclamide to block K(ATP) channel activity. Dilatations were unchanged with XAC (3.0 +/- 0.5, 3.9 +/- 0.7 and 6.1 +/- 1.0 microm), but were significantly reduced with LNNA (to 1.8 +/- 0.6, 3.5 +/- 0.7 and 4.9 +/- 0.7 microm) or glibenclamide (to 0.4 +/- 0.3, 0.8 +/- 0.7 and 1.9 +/- 0.6 microm). Neither K(ATP) channel activity nor nitric oxide was required for transmission or manifestation of the dilator response. Thus, muscle contraction can be sensed by capillaries and the signalling mechanism for the ensuing remote dilatation depends on K(ATP) channel activity and on NO, but not adenosine. Local application of 10(-4) M adenosine, 10(-4) M sodium nitroprusside or 10(-5) M pinacidil directly to capillaries initiated remote arteriolar dilatations. Thus, capillaries can respond directly to known mediators of metabolic vasodilatation, but these signalling pathways are not invariably implicated in the response to muscle contraction.

ATP-Binding Cassette Transporters↗

PubMed2002

Multiple dilator pathways in skeletal muscle contraction-induced arteriolar dilations.

To determine whether nitric oxide (NO), adenosine (Ado) receptors, or ATP-sensitive potassium (K(ATP)) channels play a role in arteriolar dilations induced by muscle contraction, we used a cremaster preparation in anesthetized hamsters in which we stimulated four to five muscle fibers lying perpendicular to a transverse arteriole (maximal diameter approximately 35-65 microm). The diameter of the arteriole at the site of overlap of the stimulated muscle fibers (the local site) and at a remote site approximately 1,000 microm upstream (the upstream site) was measured before, during, and after muscle contraction. Two minutes of 4-Hz muscle stimulation (5-15 V, 0.4 ms) produced local and upstream dilations of 19 +/- 1 and 10 +/- 1 microm, respectively. N(omega)-nitro-L-arginine (10(-4) M; NO synthase inhibitor), xanthine amine congener (XAC; 10(-6) M; Ado A(1), A(2A), and A(2B) receptor antagonist), or glibenclamide (Glib; 10(-5) M; K(ATP) channel inhibitor) superfused over the preparation attenuated the local dilation (by 29.7 +/- 12.7, 61.8 +/- 9.0, and 51.9 +/- 14.9%, respectively), but only XAC and Glib attenuated the upstream dilation (by 68.9 +/- 6.8 and 89.1 +/- 6.4%, respectively). Furthermore, only Glib, when applied to the upstream site directly, attenuated the upstream dilation (48.1 +/- 9.1%). Neither XAC nor Glib applied directly to the arteriole between the local and the upstream sites had an effect on the magnitude of the upstream dilation. We conclude that NO, Ado receptors, and K(ATP) channels are involved in the local dilation initiated by contracting muscle and that both K(ATP) channels and Ado receptor stimulation, but not NO, play a role in the manifestation of the dilation at the upstream site.

Adenosine↗

PubMedDate not supplied

Regulation of leukocyte recruitment by local wall shear rate and leukocyte delivery.

OBJECTIVE: To determine the influence of wall shear rate and leukocyte delivery on leukocyte margination, rolling, and adhesion in post-capillary venules. METHODS: Leukocyte-endothelial cell interactions were characterized in cremaster muscle venules of anesthetized mice with video microscopy under control conditions and after 3 h exposure to TNF-alpha. Hemodynamic parameters were measured with fluorescent particle tracking using confocal microscopy. RESULTS: Leukocyte recruitment to the vessel wall and leukocyte rolling increased as a function of wall shear rate (P < 0.05) over the range observed (0-200 s(-1)) in intact post capillary venules. Leukocyte delivery affected recruitment and rolling flux only in activated vessels (P < 0.05). In addition, leukocyte firm adhesion was independent of both wall shear rate and leukocyte delivery, but showed an overall increase in TNF-alpha stimulated tissues (0.9 +/- 0.2 vs. 2.7 +/- 0.6 cells/50 microm). CONCLUSIONS: Leukocytes are delivered to venules in excess of the capacity of the local endothelium to support interactions. Elevated shear forces increase leukocyte recruitment to the vessel wall, which correlates to elevated rolling flux. In contrast, leukocyte firm adhesion is primarily affected by the activation state of the tissue and not by hemodynamic factors. Overall, the capacity of endothelial cells to support leukocyte interactions primarily regulates leukocyte recruitment and is not limited by leukocyte supply.

Animals↗