Introduction

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Local treatment of UC with lidocaine has been shown to result in good symptomatic relief and restored mucosal integrity (Björck et al., 1993). Ropivacaine, a new local anesthetic currently under investigation for the treatment of UC, appears to improve inflammatory endoscopic scores and to decrease clinical symptoms after only 2 weeks of treatment (Arlander et al., 1996). Furthermore, ropivacaine has protective effects in a rat colitis model (T. Martinsson, unpublished observation).

In addition to reversible block of nerve impulse propagation, local anesthetics are known to affect a variety of other cell functions (Hammer et al., 1985; Moudgil et al., 1977; Dickstein et al., 1985). Many of these effects are related to leukocyte function; local anesthetics have been shown to inhibit leukocyte phagocytic activity (Cullen and Haschke, 1974), superoxide production (Peck et al., 1985; Irita et al., 1986) and adhesion (Giddon and Lindhe, 1972; Rabinovitch and DeStefano, 1974; MacGregor et al., 1980). Lidocaine also reduces leukocyte adherence in injured venules in vivo (Stewart et al., 1974), counteracts the endothelial damage induced by sticking leukocytes (Stewart et al., 1974) and inhibits granulocyte recruitment to sites of inflammation (MacGregor et al., 1980). Ropivacaine itself has recently been shown to inhibit release of LTB₄ and 5-hydroxyeicosatetraenoic acid from leukocytes (Martinsson et al., 1997).

The migration of leukocytes into tissues is a crucial event in the inflammatory response. Leukocyte emigration is normally responsible for the successful host response to tissue injury and infection, but it is also potentially harmful and contributes to the pathology of different inflammatory diseases such as UC (Babbs, 1992). Accordingly, it has been shown that suppression of neutrophil function may reduce tissue damage in inflammatory diseases (Fujita et al., 1994), including UC (Palmen et al., 1995).

The accumulation of leukocytes in inflamed tissue and the excessive filtration of fluid and proteins that accompanies an inflammatory response are largely confined to small venules in the microcirculation (Granger and Kubes, 1994). Leukocyte extravasation is initiated by interactions between circulating leukocytes and activated endothelial cells lining the venules of inflamed tissue. Rolling of leukocytes along the venular wall is the earliest visible interaction. This is a reversible event that can be followed by either the release of the leukocytes back into the bloodstream or, upon chemotactic stimulation, by arrest and firm adhesion to the endothelium and subsequent diapedesis (Granger and Kubes, 1994). Recent studies have revealed that the leukocyte-endothelial interactions are mediated by different classes of cell surface adhesion molecules. These include the selectins, the integrins and members of the immunoglobulin superfamily (Springer, 1994). The selectins (E-, L- and P-selectin) are required for leukocyte rolling along the vessel wall (Ley and Tedder, 1995). L-selectin is constitutively expressed on leukocytes, whereas E- and P-selectin are inducible endothelial molecules (Springer, 1994). CD11b/CD18, a member of the integrin family, appears to be important for the firm adhesion of leukocytes to the endothelium (Arfors et al., 1987). This receptor is constitutively expressed on the surface of nonactivated leukocytes, and the surface expression of CD11b/CD18 may be further increased by mobilization from intracellular pools upon stimulation with various inflammatory stimuli (Todd et al., 1984).

Two important questions arise from the previous observations about the effects of local anesthetics on leukocyte activation and the promising results of ropivacaine in the treatment of UC. First, can ropivacaine inhibit leukocyte adhesion in vivo and/or the increased vascular permeability associated with inflammation? Second, can ropivacaine affect the expression of adhesion molecules on the surface of leukocytes? In the present study, we examined the effects of ropivacaine on increased vascular permeability and inflammatory leukocyte behavior in the hamster cheek pouch microvasculature by using intravital microscopy, and we examined its effects on the expression of adhesion molecules on human leukocytes in vitro by using flow cytometry.

	Materials and Methods

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Intravital Microscopy

Male golden hamsters (90-120 g, Harlan-CPB, Austerlitz, Netherlands) were used. The cheek pouch of sodium pentobarbital-anesthetized animals was prepared for intravital microscopy according to Duling (1973) with modifications by Svensjö (1978) and Erlansson et al. (1989). The exposed cheek pouch was superfused with a bicarbonate-buffered salt solution (35°C) that was continuously equilibrated with 5% CO₂ in N₂ to maintain low oxygen tension (~4 kPa) and pH 7.35. A catheter in the left femoral vein was used for infusion of FITC-labeled dextran (MW 150,000, Bioflor HB, Uppsala, Sweden) or rhodamine (Sigma, St. Louis, MO). After positioning of the cheek pouch under the microscope (Axioscope, Zeiss), a 30-min equilibration period preceded the experiments. Rhodamine-labeled leukocytes in venules were visualized with fluorescent light epi-illumination using a ×25 water immersion lens (NA 0.60). The microscopic images were televised (Sony Trinitron, Sony CCD camera) and recorded with a Sony U-matic Video Tape Recorder for subsequent off-line analysis. In another set of experiments, nonlabeled leukocytes in postcapillary venules were studied with ordinary light transillumination using a ×40 water immersion lens (NA 0.90).

Vascular permeability. FITC-labeled dextran was injected i.v. (25 mg/100 g b.wt.) as a macromolecular tracer. The increase in microvascular permeability for large molecules was quantified by counting fluorescent leakage sites at postcapillary venules (Erlansson et al., 1989). The number of leaks per square centimeter of cheek pouch area was counted before and during 30 min after topical application of LTB₄ (10 nM; Sigma; repeated four times with 45-min intervals). Ropivacaine (Naropin, Astra, Södertälje, Sweden) was applied to the superfusate for 15 min, starting 10 min before the second (10 µM ropivacaine) and the fourth (100 µM ropivacaine) LTB₄ applications. The peak number of leakage sites, which consistently occurred at 5 min after the start of LTB₄ application, was used for statistical calculations.

Leukocyte behavior. The cheek pouches were subjected to four repeated local applications of 10 nM LTB₄ for 5 min with 45- to 60-min intervals. Before the second and fourth LTB₄ applications, 100 µM ropivacaine was applied locally for 15 min starting 10 min before LTB₄; this resulted in two experimental series termed first and second. Venular segments with a diameter of 40 µm and a length of 150 µm were selected for observation of free-flowing, rolling and adherent leukocytes. All systemic leukocytes were labeled in vivo by an i.v. injection of rhodamine (2 µg) immediately before observations. The weak red fluorescence of rhodamine-labeled leukocytes was amplified using a Hamamatsu Image Intensifier and recorded for subsequent off-line analysis. Values for free-flowing, rolling and firmly adherent leukocytes were obtained immediately before LTB₄ application and during and after ropivacaine and/or LTB₄ application. Free-flowing leukocytes (with the same velocity as erythrocytes) were determined by counting the number of leukocytes passing a line perpendicular to the vessel per minute. Rolling leukocytes were defined as leukocytes that were in contact with the venular endothelium and had a velocity lower than that of free-flowing leukocytes, and the rolling leukocyte flux (cells per minute) was determined as described for free-flowing leukocytes. The leukocyte rolling fraction was determined at indicated time-points by dividing the rolling leukocyte flux by the total leukocyte flux (flux of rolling leukocytes plus that of free-flowing leukocytes). Cells were considered to be adherent if they remained stationary for more than 30 sec. Adherent cells were expressed as number of leukocytes/10,000 µm² inner surface of the vessel. In a second set of experiments, postcapillary venules with a diameter of 10 µm and a length of 100 µm were selected for observation of leukocytes with ordinary light transillumination. In these experiments, rolling leukocyte flux and adherent leukocytes were counted.

Vessel diameters. The diameters of venules and arterioles were measured off-line using an image-shearing monitor (IPM, LaMesa, CA).

Expression of Cell Surface Adhesion Molecules

Preparation of leukocytes. EDTA blood from healthy human donors (n = 19) was hemolyzed by dilution (1:20) in 4°C isotonic NH₄Cl-EDTA lysing solution (154 mM NH₄Cl, 10 mM KHCO₄, 0.1 mM EDTA, pH 7.2). After incubation for 5 min at 15°C, the leukocytes were centrifuged (300 × g, 4°C) for 5 min. The leukocyte pellets were washed once in 4°C 0.15 M PBS supplemented with 0.1 M EDTA and 0.02% sodium azide (PBS-EDTA). This cell preparation procedure minimizes spontaneous leukocyte activation (Lundahl et al., 1991).

Leukocyte activation. The basic medium used during leukocyte activation was RPMI 1640 (Gibco Ltd., Paisley, U.K.) containing 0.01 M HEPES and 5% fetal calf serum (Gibco Ltd.). TNF- (R&D Systems, Abingdon, U.K.) was diluted to a final concentration of 10¹⁰ g/ml. Ropivacaine and lidocaine (Xylocaine, Astra) were diluted in medium to make serial dilutions ranging from 10⁵ M to 10³ M. The leukocytes were incubated with or without TNF- for 15 min (L-selectin) or 30 min (CD11b/CD18), both at 37°C. The cells were treated with varying concentrations of ropivacaine, lidocaine or an equal volume of medium, added together with TNF-. The activation was stopped by addition of cold PBS-EDTA, and the leukocytes were washed once, resuspended in 100 µl PBS-EDTA and kept on ice. As a control of spontaneous cell activation at 37°C, leukocytes were also incubated at 4°C without TNF- and local anesthetics. The viability of the cells before and after incubation with ropivacaine or lidocaine was >95%, as determined by the trypan blue exclusion test.

Flow cytometric analysis of adhesion molecule expression. The expression of CD11b and L-selectin on granulocytes and monocytes was analyzed by adding 5 µl of PE-conjugated monoclonal anti-CD11b (DAKO A/S, Glostrup, Denmark) or 10 µl FITC-conjugated anti-L-selectin antibody (Becton Dickinson, Mountain View, CA) to the leukocytes prepared and treated as described above. The suspensions were incubated on ice for 30 min, washed in cold PBS-EDTA and resuspended in 0.5 ml of cold PBS-EDTA before analysis. Appropriate concentrations of iso- and subtype-matched control antibodies were used to define the cutoff for positive fluorescence. Positive fluorescence was the 99th percentile of the distribution of the cells labeled with the respective control antibody (PE-conjugated IgG_2a and FITC-conjugated IgG_2a for CD11b and L-selectin, respectively).

Intracellular Free Ca⁺⁺ Concentration

Leukocytes were prepared as described previously. Intracellular free Ca⁺⁺ concentration was measured as described by Tuominen et al. (1994). Briefly, the cells were loaded with 2.5 µM fura-2 AM in Ca⁺⁺- and Mg⁺⁺-free bufferHanks' Balanced Salt Solution (Gibco Ltd.) supplemented with 10 mM glucoseat 37°C for 30 min. The cells were washed once (440 × g, 5 min) and resuspended in buffer (10⁶ cells/ml) and stored at room temperature. Aliquots of 2 × 10⁶ cells were diluted in Ca⁺⁺-containing (1 mM) buffer, and fluorescence was recorded at 340/380 nm (excitation) and 510 nm (emission) in a Perkin Elmer spectrofluorometer equipped with a thermostatically controlled cuvette holder (37°C) and constant stirring. The dye response was calibrated by the sequential addition of 10 µM ionomycin and 50 mM EGTA at the end of the experiment to give the maximum and minimum fluoresence, respectively.

Statistical methods. Mean values and standard errors (S.E.M.) were calculated. The results were tested by analysis of variance (ANOVA) followed by Student's t test for paired comparisons. P < .05 was considered significant.

	Results

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Leukocyte expression of CD11b/CD18. The granulocytes incubated at 37°C were spontaneously activated to some extent; that is, the expression of CD11b/CD18 on the cell surface was increased 3-fold, measured as increased MFI values (from 9 ± 1.4 to 36 ± 3.4). TNF- stimulation further increased expression of CD11b/CD18 by 80% (65 ± 2.4, P < .001). Ropivacaine 100 µM and lidocaine 300 µM significantly reduced this up-regulation, whereas the lower concentrations were inactive in this regard (fig. 1). The TNF-- induced CD11b/CD18 up-regulation was abolished by the local anesthetics at 1 mM. The expression of CD11b/CD18 on monocytes after activation with TNF- was lower than the expression on granulocytes; however, the effect mediated by the local anesthetics was similar to that described above (data not shown). The 100 µM concentration of ropivacaine was chosen for the in vivo experiments because this was the lowest concentration with significant effects on CD11b/CD18 expression. This concentration is also in the therapeutic range obtained in the colon of patients treated rectally with ropivacaine (Arlander et al., 1996).

View larger version (18K): [in this window] [in a new window]   Fig. 1.   Dose-response curves of the cell surface expression of CD11b/CD18 on human granulocytes (n = 10 for ropivacaine and n = 9 for lidocaine). Cells were incubated with TNF- (1010 g/ml) and ropivacaine () or lidocaine () for 30 min at 37°C. Incubation with TNF- alone was used as control. Ropivacaine and lidocaine dose-dependently reduced CD11b/CD18 up-regulation. Values are means ± S.E.M. *P < .05, **P < .01, ***P < .001.

Leukocyte adhesion. Figure 2 shows the number of adherent cells in venules (ø = 40 µm) of the hamster cheek pouch at different time-points, together with the rolling leukocyte flux. The base-line venular leukocyte adhesion was 10.2 ± 3.6 cells/10,000 µm². Superfusion with LTB₄ for 5 min significantly increased the number of adherent cells by 78%. This increase was reversible and returned to base line after termination of LTB₄ application. Addition of ropivacaine to the superfusion solution significantly reduced the leukocyte adhesion response to LTB₄ to a value comparable to the base-line value (fig. 2; table 1). Ropivacaine showed a tendency to reduce spontaneous adhesion, but this effect was not significant. The results could be repeated in the second series of experiments in the same preparation. In this second series, LTB₄ increased leukocyte adhesion by 115% (compared with 78% in the first LTB₄ application), and ropivacaine inhibited the induced adhesion completely (data not shown).

View larger version (15K): [in this window] [in a new window]   Fig. 2.   Number of adherent (panel A) and rolling (panel B) leukocytes in venules of the hamster cheek pouch at indicated time-points. LTB4 (10 nM) was applied topically for 5 min, and ropivacaine (100 µM) was applied for 15 min starting 10 min before LTB4 application. Ropivacaine significantly (P < .05) inhibited the LTB4-induced adhesion and also by itself inhibited the spontaneous rolling leukocyte flux (P < .05). Data points and error bars represent means ± S.E.M.

Leukocyte rolling. The base-line rolling leukocyte flux in venules was 26.6 ± 6.9 cells/min (table 1). As a result of the increased adhesion induced by LTB₄, rolling decreased markedly (94%) during the 5-min LTB₄ application (fig. 2). Ropivacaine reduced the spontaneous rolling flux by 72% without causing increased adhesion (fig. 2; table 1). During the combined application of ropivacaine and LTB₄, the rolling leukocyte flux was further reduced (76%) despite the inhibition of LTB₄-induced adhesion during this time. The leukocyte rolling fraction was calculated, and during base-line conditions, the fraction of rolling cells was 41% ± 4.2% (table 1). Ropivacaine reduced the rolling fraction by half. In the second series in the same preparations, ropivacaine significantly reduced the spontaneous leukocyte rolling flux and the rolling fraction by 57% and 47%, respectively (data not shown).

Total flux of leukocytes. The total number of free-flowing and rolling leukocytes, a value that reflects blood flow, is shown in table 1. The base-line total leukocyte flux was 61.8 ± 11.6 cells/min, and ropivacaine reduced the total flux by 47%. Ropivacaine significantly reduced the total leukocyte flux by 35% on repetition of the experiments in the same preparation (data not shown).

Vessel diameters. Ropivacaine (100 µM) applied topically for 15 min significantly reduced arteriolar diameters by 43% ± 4.4% (P < .05, n = 6), whereas venular diameters were not significantly affected.

Vascular permeability. In line with previous observations (Erlansson et al., 1989), topical LTB₄ challenge in the hamster cheek pouch resulted in reversible increases in the number of postcapillary leakage sites (plasma exudation). When 10 or 100 µM ropivacaine was applied topically before LTB₄, the peak number of leakage sites was markedly reduced (fig. 3). The effect was fully reversible on washout after 10 µM ropivacaine, but it was only partially reversible after 100 µM.

View larger version (16K): [in this window] [in a new window]   Fig. 3.   Number of postcapillary leakage sites per square centimeter after five repeated applications of 10 nM LTB4 for 5 min to the hamster cheek pouch. The first application of LTB4 greatly increased permeability (1). Starting 10 min before the second and fourth LTB4 applications, ropivacaine (black bars) was added to the superfusate at a final concentration of 10 µM (2) and 100 µM (4), respectively. The ropivacaine-mediated inhibition was reversible as shown by LTB4 challenge during the washout periods (3) and (5). The results are mean values ± S.E.M. in six hamsters. **P < .01, ***P < .001 as compared with the first LTB4 application (1).

Leukocyte expression of L-selectin. L-selectin is rapidly shed by proteolytic cleavage after leukocyte activation (Kishimoto et al., 1989). Accordingly, the granulocyte membrane expression of L-selectin was decreased by 50% (from 13 ± 1.8 to 6.5 ± 0.3) after 15 min of activation (TNF-) as compared with controls. Ropivacaine and lidocaine dose-dependently suppressed L-selectin shedding, the lowest concentrations with significant effects being 100 and 300 µM, respectively, (fig. 4). In addition, ropivacaine inhibited the shedding of L-selectin on monocytes. However, this effect was not so pronounced as that seen for granulocytes (data not shown). Lidocaine was inactive in this regard.

View larger version (18K): [in this window] [in a new window]   Fig. 4.   Dose-response curves of L-selectin expression on human granulocytes (n = 6 for ropivacaine and n = 8 for lidocaine). Cells were incubated with TNF- (1010 g/ml) and with ropivacaine () or lidocaine (), respectively, for 15 min at 37°C. Incubation with TNF- alone was used as control. Ropivacaine and lidocaine dose-dependently inhibited L-selectin shedding. Values are means ± S.E.M. *P < .05, **P < .01.

Intracellular Ca⁺⁺ concentrations. Stimulation of leukocytes with LTB₄ (0.01, 0.1 and 1 µM) induced a dose-dependent rise in [Ca⁺⁺]_ifrom a basal level of 115 nM to 290, 350 and 405 nM, respectively (n = 4). Pretreatment of the leukocytes for 10 min with 1 mM ropivacaine did not affect the LTB₄-induced Ca⁺⁺ transients significantly, i.e., from 175 nM to 350, 400, and 465 nM, respectively.

	Discussion

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A key event in inflammation is the recruitment of leukocytes to sites of inflammation. This recruitment consists of several sequential steps, including leukocyte rolling along the endothelium followed by firm adhesion of the leukocytes to the endothelial cells. In the present study, we show that ropivacaine can reduce both the rolling and the firm adhesion in vivo, as well as the increased vascular permeability associated with leukocyte adhesion. Furthermore, ropivacaine was found to inhibit the induced expression of CD11b/CD18 on leukocytes in vitro.

Ropivacaine almost completely inhibited the LTB₄-induced leukocyte adhesion in both postcapillary and larger venules. This may have been the result of a direct effect on firm adhesion and/or, given that venular rolling is a prerequisite for firm adhesion (Lindbom et al., 1992), an indirect effect mediated through inhibition of leukocyte rolling. Because LTB₄ has been shown to stimulate leukocyte adhesion through CD11b/CD18 (Arfors et al., 1987; Tonnesen et al., 1989), our in vitro finding that ropivacaine inhibited up-regulation of neutrophil CD11b/CD18 indicates that the observed effect of ropivacaine on adhesion was, at least in part, mediated through reduction of CD11b/CD18 expression. Interestingly, it has been suggested that CD11b/CD18 is involved in the interactions between intestinal epithelial cells and neutrophils (Parkos et al., 1995), which implies that ropivacaine may also interfere with transepithelial leukocyte migration.

The mechanism by which the local anesthetics inhibited expression of CD11b/CD18 is unknown. However, because local anesthetics are known to act on ion channels to decrease membrane permeability to Na⁺ and K⁺ in nerves and may have a similar action on other cell types, one possibility is that local anesthetics interact with different ion channels on the leukocytes. Leonard et al. (1992) have found that the membrane potential of resting T cells is set by voltage-activated channels and that blockage of these channels is sufficient to depolarize resting human T cells and prevent their activation. Thus, if the membrane potential is part of the leukocyte activation system, the underlying mechanism for the observed inhibition of CD11b/CD18 expression by local anesthetics could be explained in these terms. Another mechanism by which ropivacaine could inhibit the expression of CD11b/CD18 is through the inhibition of LTB₄ signal transduction, e.g., by an action on calcium channels or stores. However, the results so far do not support a hypothesis of an action on leukocyte calcium mechanisms. It could also be speculated that the effect on adhesion is due to inhibition of endogenous LTB₄. This is unlikely, however, because 5-lipoxygenase inhibitors have been shown to be ineffective in acute (as in this study) leukocyte-dependent LTB₄-induced responses in the hamster cheek pouch (Raud, 1989).

The close relationship between initial leukocyte rolling flux and subsequent adhesion (Lindbom et al., 1992; Mayrovitz, 1992) suggests another mechanism by which ropivacaine might inhibit adhesion: inhibition of leukocyte rolling. We found that ropivacaine reduced venular rolling leukocyte flux in vivo by 70%, and, in contrast to the LTB₄-mediated effect, the effect of ropivacaine was not due to increased adhesion. Inhibition of the rolling flux may be the result of a reduced fraction of rolling leukocytes and/or reduced delivery of leukocytes (i.e., blood flow). Ropivacaine was found to inhibit the rolling leukocyte fraction by approximately 50% compared with the control condition. Because the magnitude of the rolling leukocyte fraction is dependent on the selectins and/or their ligands (Ley and Tedder, 1995), the latter finding indicates that ropivacaine somehow interfered with selectin expression or function. It has been demonstrated that the spontaneous leukocyte rolling observed after preparation of tissues for intravital microscopy is mediated by both L- and P-selectin (Todd et al., 1984; Doré, et al., 1993; von Andrian et al., 1991). However, it is unlikely that the inhibitory effect of ropivacaine on the leukocyte rolling was related to L-selectin expression, because treatment with 100 µM ropivacaine retained surface expression of L-selectin on the granulocytes. This leaves endothelial P-selectin as a possible target of ropivacaine. Furthermore, local anesthetics have been suggested to "stabilize" the cell membrane of leukocytes (Young and MacKenzie, 1992), and L-selectin and the P-selectin glycoprotein ligand-1 are localized on the microvilli of neutrophils to improve the presentation of these molecules to the endothelium (Patel et al., 1995). Therefore, it is possible that ropivacaine reduces rolling by changing leukocyte cell membrane morphology and adhesion molecule distribution.

The inhibition of the rolling leukocyte fraction accounted for approximately 50% of the effect of ropivacaine on the rolling leukocyte flux. The remaining effect by ropivacaine on the rolling flux appeared to be related to a reduction in blood flow, detected as a partial arteriolar constriction and as a significant reduction in the total leukocyte flux (which reflects blood flow), a value that is known to be correlated with the rolling leukocyte flux (Thorlacius et al., 1995). We thus suggest that the inhibitory effect by ropivacaine on leukocyte rolling was partly due to changes in leukocyte-endothelium adhesive interactions and partly related to alterations in blood flow.

Ropivacaine markedly inhibited the LTB₄-induced plasma leakage in a dose-dependent and reversible manner. Because LTB₄-induced plasma extravasation is mediated by leukocytes (Björk et al., 1982; Kurose et al., 1994), the inhibition of plasma leakage by ropivacaine may be a result of its ability to reduce leukocyte-endothelial cell interactions.

The inhibitory effect of ropivacaine on leukocyte adhesion differs from the anti-inflammatory action of glucocorticoids, which are commonly used for the local treatment of UC. In contrast to ropivacaine, glucocorticoids do not inhibit the increased endothelial adhesion induced by chemotactic factors but instead inhibit the leukocyte extravasation process (Oda and Katori, 1992). Interestingly, metronidazole, a potent antimicrobial agent that is gaining recognition as a possible mode of therapy for treatment of UC, has effects comparable to those of ropivacaine. This agent has been shown to inhibit LTB₄-induced adhesion in the microcirculation of the rat mesentery (Arndt et al., 1994). With regard to lidocaine, another drug tested for treatment of UC, our study confirms previous observations that this local anesthetic can inhibit CD11b/CD18 up-regulation and L-selectin down-regulation on neutrophils (Ohsaka et al., 1994). However, we found that lidocaine was 2.5 times less potent than ropivacaine in inhibiting the CD11b/CD18 expression.

In conclusion, ropivacaine was found to inhibit inflammatory leukocyte rolling, firm adhesion and the associated increased vascular permeability in vivo. Moreover, our in vitro findings showed that ropivacaine had an inhibitory effect on induced expression of CD11b/CD18. Because leukocyte-endothelial cell interactions represent early and rate-limiting steps in intestinal inflammatory processes, these findings may help explain the beneficial effect of ropivacaine seen in the treatment of UC.