Effects of different leukocyte subpopulations and flow conditions on leukocyte accumulation during reperfusion.

BACKGROUND/AIMS: The study examined the interdependent effects of shear stress and different leukocyte subpopulations on endothelial cell activation and cell interactions during low flow and reperfusion. METHODS: Human umbilical venous endothelial cells were perfused with either neutrophils or monocytes at different shear stress (2-0.25 dyn/cm(2)) and adhesion was quantified by microscopy. Effects of adherent neutrophils and monocytes on endothelial cell adhesion molecule expression were analyzed by flow cytometry after 4-hour static coincubation. After coincubation, the cocultures were reperfused with labeled neutrophils at 2 dyn/cm(2) and their adhesion was quantified selectively. For the control, endothelium monocultures with and without lipopolysaccharide activation were used. RESULTS: At 2 dyn/cm(2), adhesion did not exceed baseline levels on nonactivated endothelium. Decreasing shear stress to 0.25 dyn/cm(2) largely increased the adhesion of both leukocyte subpopulations, similar to the effect of lipopolysaccharide at 2 dyn/cm(2). However, only adherent monocytes increased adhesion molecule expression, whereas neutrophils had no effect. As a functional consequence, adherent monocytes largely increased neutrophil adhesion during reperfusion, whereas adherent neutrophils did not. CONCLUSION: Compromised shear stress is an autonomous trigger of leukocyte adhesion even in the absence of additional activators. Exceeding this immediate effect, adherent monocytes induce further endothelial activation and enhance further neutrophil adhesion during reperfusion.

ImageStream cytometry extends the analysis of phagocytosis and oxidative burst.

AIM: Phagocytosis is often measured using conventional microscopy and flow cytometry. ImageStream cytometry is a new technology that combines the advantages of both methods, enabling statistically robust microscopic applications. We compared ImageStream cytometry to flow cytometry in a whole blood model of phagocytosis with viable, fluorescence-marked Staphylococcus aureus. We furthermore measured the co-localization of intracellular bacteria to sites of oxidative burst, as well as changes in cell size and actin levels as a result of phagocytosis. EXPERIMENTAL DESIGN: Fluorescence-labeled S. aureus in a ratio of 5:1 bacteria per leukocyte were added to whole blood. Phagocytosis was stopped at different time points. After staining of neutrophils and lysis of erythrocytes, samples were analysed by ImageStream cytometry and flow cytometry. RESULTS: Phagocytosis and oxidative burst determined by flow cytometry and ImageStream cytometry showed strong correlation. In contrast to flow cytometry, ImageStream cytometry easily detected and excluded extracellular adherent bacteria from the measurement of phagocytosis, and enumerated the bacteria within each neutrophil. Using the Bright Detail Similarity score, we identified a subset of neutrophils with intracellular bacteria co-localized to sites of oxidative burst activity. Phagocytosis resulted in an increase in cell size and actin polymerization as determined by an increase in phalloidin fluorescence intensity. CONCLUSIONS: We describe a simple whole blood image-based method for measuring bacterial phagocytosis and oxidative burst. ImageStream cytometry provides the spatial resolution to determine the number of bacteria ingested and the sub-cellular localization and trafficking patterns that enables a more complete evaluation of the phagocytic process.

Mechanisms of leukocyte distribution during sepsis: an experimental study on the interdependence of cell activation, shear stress and endothelial injury.

INTRODUCTION: This study was carried out to determine whether interactions of cell activation, shear stress and platelets at sites of endothelial injury explain the paradoxical maldistribution of activated leukocytes during sepsis away from local sites of infection towards disseminated leukocyte accumulation at remote sites. METHODS: Human umbilical venous endothelial cells (HUVEC) and polymorphonuclear neutrophils (PMN) were activated with lipopolysaccharide at 100 and 10 ng/ml to achieve adhesion molecule patterns as have been reported from the hyper- and hypo-inflammatory stage of sepsis. To examine effects of leukocyte activation on leukocyte-endothelial interactions, activated HUVEC were perfused with activated and non-activated neutrophils in a parallel plate flow chamber. Adhesion molecule expression and function were assessed by flow cytometry and blocking antibodies. In a subset of experiments the sub-endothelial matrix was exposed and covered with platelets to account for the effects of endothelial injury. To investigate interactions of these effects with flow, all experiments were done at various shear stress levels (3 to 0.25 dyne/cm(2)). Leukocyte-endothelial interactions were analyzed by videomicroscopy and analysis of covariance. RESULTS: Activation of neutrophils rendered adhesion increasingly dependent on shear stress reduction. At normal shear stress, shedding of L-selectin decreased adhesion by 56%. Increased rolling fractions of activated PMN at low shear stress revealed impaired integrin affinity despite numerical up-regulation of CD11b. On sub-maximally activated, intact HUVEC shear stress became the prevailing determinant of adhesion. Presence of a platelet-covered injury with high surface density of P-selectin was the strongest variable for adhesion. When compared to maximally activated HUVEC, platelets increased neutrophil adhesion by 2.7-fold. At sub-maximal activation a 10-fold increase was observed (P < 0.05 for all). CONCLUSIONS: L-selectin shedding and integrin dysfunction render leukocyte adhesion increasingly susceptible to shear stress and alternative adhesion receptors. In combination, these effects inhibit recruitment to normally perfused sites with intact endothelium and favor maldistribution towards sites with compromised perfusion or endothelial injury.

Systemic lidocaine decreased the perioperative opioid analgesic requirements but failed to reduce discharge time after ambulatory surgery.

BACKGROUND: In this randomized, blinded, placebo-controlled trial, we evaluated whether systemic lidocaine would reduce pain and time to discharge in ambulatory surgery patients. METHODS: Sixty-seven patients were enrolled to receive lidocaine or saline infusion perioperatively. RESULTS: Length of postanesthesia care unit (PACU) stay did not differ between groups. Intraoperative opioid use was significantly less in the lidocaine group, both in the PACU and during the total study period but not after discharge. In the PACU, patients in the lidocaine group reported less pain (visual analog scale score 3.1 +/- 2.04 vs 4.5 +/- 2.9; P = 0.043). There were no differences in postoperative nausea and vomiting. CONCLUSION: Perioperative systemic lidocaine significantly reduces opioid requirements in the ambulatory setting without affecting time to discharge.

The inhibition of human neutrophil phagocytosis and oxidative burst by tricyclic antidepressants.

BACKGROUND: Tricyclic antidepressants are being investigated as long-acting analgesics for topical application in wounds or IV for postoperative pain relief. However, it remains unclear if tricyclic antidepressants affect the host defense and if reported toxic effects on neutrophils are of relevance in this setting. We therefore investigated the effects of amitriptyline, nortriptyline, and fluoxetine on human neutrophil phagocytosis, oxidative burst, and neutrophil toxicity in a human whole blood model. METHODS: Heparinized blood samples from healthy volunteers were incubated with amitriptyline, nortriptyline, or fluoxetine (10(-6) to 10(-3) M) for 0, 1, or 3 h. Staphylococcus aureus in a bacteria:neutrophil ratio of 5:1 and dihydroethidium (for the determination of oxidative burst) were added. Phagocytosis was stopped after 5, 10, 20, and 40 min. After lysis of red blood cells, samples were analyzed by flow cytometry. RESULTS: In concentrations up to 10(-4) M, none of the compounds affected neutrophil phagocytosis and oxidative burst. At 10(-3) M, all three compounds were highly toxic for neutrophils. Amitriptyline preserved morphological integrity, but completely suppressed neutrophil function. Nortriptyline and fluoxetine caused a marked disruption of neutrophils. The effects of the investigated antidepressants were not time-dependent. CONCLUSIONS: Phagocytosis and intracellular host defense are largely unaffected by antidepressants in concentrations of 10(-4) M and below. Our results confirm that antidepressants are highly toxic to neutrophils in millimolar concentrations. The neurotoxic effects and clinical side effects, but not effects on neutrophil functions, therefore, are likely to be the limiting factors in using antidepressants as analgesics.

Efficacy of ketamine in anesthetic dosage for the treatment of refractory complex regional pain syndrome: an open-label phase II study.

OBJECTIVE: Advanced complex regional pain syndrome (CRPS) remains very difficult to treat. While subanesthetic low-dose ketamine has shown promise in early localized CRPS, its use in advanced CRPS has not been as effective. Since ketamine's analgesic potency and duration of effect in neuropathic pain are directly dose-dependant, we investigated the efficacy of ketamine in anesthetic dosage in refractory CRPS patients that had failed available standard therapies. METHODS: Twenty ASA I-III patients suffering from refractory CRPS received ketamine in anesthetic dosage over 5 days. Outcome criteria were pain relief, effect on the movement disorder, quality of life, and ability to work at baseline and up to 6 months following treatment. RESULTS: Significant pain relief was observed at 1, 3, and 6 months following treatment (93.5 +/- 11.1%, 89.4 +/- 17.0%, 79.3 +/- 25.3%; P < 0.001). Complete remission from CRPS was observed at 1 month in all patients, at 3 months in 17, and at 6 months in 16 patients. If relapse occurred, significant pain relief was still attained at 3 and 6 months (59.0 +/- 14.7%, P < 0.004; 50.2 +/- 10.6%, P < 0.002). Quality of life, the associated movement disorder, and the ability to work significantly improved in the majority of patients at 3 and 6 months. CONCLUSIONS: This open-label trial suggests benefit in pain reduction, associated CRPS symptoms, improved quality of life and ability to work following anesthetic ketamine in previously refractory CRPS patients. However, a randomized controlled trial will be necessary to prove its efficacy.

Complete recovery from intractable complex regional pain syndrome, CRPS-type I, following anesthetic ketamine and midazolam.

OBJECTIVE: To describe the treatment of an intractable complex regional pain syndrome I (CRPS-I) patient with anesthetic doses of ketamine supplemented with midazolam. METHODS: A patient presented with a rapidly progressing contiguous spread of CRPS from a severe ligamentous wrist injury. Standard pharmacological and interventional therapy successively failed to halt the spread of CRPS from the wrist to the entire right arm. Her pain was unmanageable with all standard therapy. As a last treatment option, the patient was transferred to the intensive care unit and treated on a compassionate care basis with anesthetic doses of ketamine in gradually increasing (3-5 mg/kg/h) doses in conjunction with midazolam over a period of 5 days. RESULTS: On the second day of the ketamine and midazolam infusion, edema, and discoloration began to resolve and increased spontaneous movement was noted. On day 6, symptoms completely resolved and infusions were tapered. The patient emerged from anesthesia completely free of pain and associated CRPS signs and symptoms. The patient has maintained this complete remission from CRPS for 8 years now. CONCLUSIONS: In a patient with severe spreading and refractory CRPS, a complete and long-term remission from CRPS has been obtained utilizing ketamine and midazolam in anesthetic doses. This intensive care procedure has very serious risks but no severe complications occurred. The psychiatric side effects of ketamine were successfully managed with the concomitant use of midazolam and resolved within 1 month of treatment. This case report illustrates the effectiveness and safety of high-dose ketamine in a patient with generalized, refractory CRPS.

"Imagine your neighbor mows the lawn": a pilot study of psychological sequelae due to awake craniotomy: clinical article.

OBJECT: Although it has been reported that awake neurosurgical procedures are well tolerated, the long-term occurrence of general psychological sequelae has not yet been investigated. This study assessed the frequency and effects of psychological symptoms after an awake craniotomy on health-related quality of life (HRQOL). METHODS: Sixteen patients undergoing an awake surgery were surveyed with a self-developed questionnaire, the Posttraumatic Stress Disorder Inventory For Awake Surgery Patients, which adopts the core components of the Diagnostic and Statistical Manual of Mental Disorders (Fourth Edition) posttraumatic stress disorder (PTSD) criteria. The mean time between surgery and data collection was 97.3 ± 93.2 weeks. Health-related quality of life was assessed with the 36-Item Short Form Health Survey. RESULTS: Forty-four percent of the patients stated that they had experienced either repetitive distressing recollections or dreams related to the awake surgery, 18.8% stated persistent avoidance of stimuli associated with the awake surgery, and symptoms of increased arousal occurred in 62.5%. Two patients presented with postoperative psychological sequelae resembling PTSD symptoms. Younger age at surgery and female sex were risk factors for symptoms of increased arousal. The experience of intense anxiety during awake surgery appears to favor the development of postsurgical PTSD symptoms, while recurrent distressing recollections particularly affect HRQOL negatively. CONCLUSIONS: In many cases awake craniotomy is necessary to preserve language and motor function. However, in some cases awake craniotomy can lead to postoperative psychological sequelae resembling PTSD symptoms. Therefore, possible long-term effects of an awake surgery should be considered and discussed with the patient when planning this type of surgery.

Local anesthetic effects on human neutrophil priming and activation.

BACKGROUND: The anti-inflammatory effects of local anesthetics (LAs) are well documented. Local anesthetics in micromolar concentrations inhibit extracellular oxygen release in isolated neutrophils; the underlying mechanism seems to be an inhibition of leukocyte priming. It remains unclear, however, if first, these effects also can be observed in whole blood, and second, if the priming of other neutrophil functions is similarly attenuated by LAs. Furthermore, the effects of LAs on intracellular generation of oxidative species remain to be investigated. METHODS: Whole-blood samples from healthy volunteers were incubated for 0, 1, or 3 hrs with different concentrations (10 to 10 M) of either lidocaine, ropivacaine, QX314, or NaCl 0.9% as control. Dihydroethidium was added to quantify oxidative burst. Samples were primed with platelet-activating factor (PAF, 10 M) and/or activated with formyl-methyl-leucyl-phenylalanine (10 M) for 15 mins each. After staining for CD11b and lysis of erythrocytes, samples were analyzed by flow cytometry. RESULTS: Priming of leukocytes is a relevant mechanism in whole blood. Platelet-activating factor stimulates the priming of oxidative burst and CD11b expression. Lidocaine up to millimolar concentrations did not affect the PAF priming and formyl-methyl-leucyl-phenylalanine activation of oxidative burst. The priming of CD11b expression and the priming and activation of changes in cell morphology were significantly attenuated by lidocaine. CONCLUSIONS: The intracellular generation of reactive oxygen species remains largely unaffected by LAs in clinical concentrations. This suggests that the anti-inflammatory effects of LAs do not interfere with the host defense.

Local anesthetics time-dependently inhibit staphylococcus aureus phagocytosis, oxidative burst and CD11b expression by human neutrophils.

BACKGROUND AND OBJECTIVES: Local anesthetics have been shown to modulate neutrophil functions in a time-dependent manner, which might help to prevent inflammatory injury to the organism. However, if host defense mechanisms are affected similarly, the ability to eliminate bacteria might be reduced. We hypothesized that local anesthetics have time-dependent effects on phagocytosis of S. aureus, oxidative burst, and CD11b expression by human neutrophils. To test this hypothesis, we reanalyzed data from a previous study. METHODS: Blood samples from 11 healthy volunteers were incubated with lidocaine (1,846 mumol/L), bupivacaine (770 mumol/L) or ropivacaine (801 mumol/L) for 30 minutes. Thereafter, bacteria were added, either fluorescently labeled for determination of phagocytosis, or unstained for determination of oxidative burst and CD11b expression. After an additional incubation for 0, 10, 30, or 60 minutes, phagocytosis was stopped and neutrophils were stained with monoclonal antibodies for flow cytometric analysis. Data were analyzed by analysis of variance for repeated measurements. RESULTS: Lidocaine and bupivacaine inhibited neutrophil functions in a time-dependent manner (P < .05). Prolonged local anesthetic exposure reduced the fraction of ingesting neutrophils by 20% +/- 12% (mean +/- SD) and 7% +/- 7%, bacterial uptake by 19% +/- 16% and 14% +/- 12%, oxidative burst by 29% +/- 23% and 28% +/- 25%, and CD11b expression by 66% +/- 24% and 25% +/- 21% for lidocaine and bupivacaine, respectively. Ropivacaine exerted a time-dependent effect on CD11b expression only (24% +/- 34%; P < .05). CONCLUSIONS: Our results indicate that in a whole blood model, time-dependent effects of local anesthetics affect key neutrophil functions necessary for bacterial elimination. However, these effects only occur at concentrations that are unlikely to be routinely attained in the clinical setting, and concern about interfering with the host defense is likely unwarranted.

A pilot open-label study of the efficacy of subanesthetic isomeric S(+)-ketamine in refractory CRPS patients.

OBJECTIVE: Complex regional pain syndrome (CRPS) is a severe neuropathic pain state that is often disproportionate to the initial trauma. Associated features are autonomic dysregulation, swelling, motor dysfunction, and trophic changes to varying degrees. Despite a multitude of treatment modalities, a subgroup of CRPS patients remain refractory to all standard therapies. In these patients, the disease may spread extraterritorially, which results in severe disability. A critical involvement of N-methyl-D-aspartate receptors (NMDARs) has been demonstrated both clinically and by animal experimentation. NMDA antagonists may be effective in many neuropathic pain states. In long-standing, generalized CRPS, we investigated the effects of S(+)-ketamine on pain relief and somatosensory features, assessed by quantitative sensory testing (QST). METHODS: Four refractory CRPS patients received continous S(+)-ketamine-infusions, gradually titrated (50 mg/day-500 mg/day) over a 10-day period. Pain intensities (average, peak, and least pain) and side effects were rated on visual analogue scales, during a 4-day baseline, over 10 treatment days, and 2 days following treatment. QST (thermo-, mechanical detection, and pain thresholds) was analyzed at baseline and following treatment. RESULTS: Subanesthetic S(+)-ketamine showed no reduction of pain and effected no change in thermo- and mechanical detection or pain thresholds. This procedure caused no relevant side effects. The lack of therapeutic response in the first four patients led to termination of this pilot study. CONCLUSION: S(+)-ketamine can be gradually titrated to large doses (500 mg/day) without clinically relevant side effects. There was no pain relief or change in QST measurements in this series of long-standing severe CRPS patients.

Dose-dependent influence of barbiturates but not of propofol on human leukocyte phagocytosis of viable Staphylococcus aureus.

OBJECTIVE: Deep sedation with barbiturates or propofol is a standard therapy for patients with critically elevated intracranial pressure. Such patients are prone to infectious complications, especially to pneumonias, which are most commonly caused by Staphylococcus aureus. Although various immunomodulatory effects of barbiturates have been described in vitro, their influence on the phagocytosis of viable S. aureus has yet to be investigated. Therefore, we examined the effects of thiopentone, methohexitone, and propofol on the phagocytosis of viable S. aureus. DESIGN: Laboratory study. SETTING: University laboratory. PATIENTS: Ten healthy volunteers aged 32.5 +/- 7 yrs. INTERVENTIONS: Blood sampling. MEASUREMENTS AND MAIN RESULTS: Whole blood samples were preincubated with different concentrations of thiopentone, methohexitone, and propofol, which is an isopropylphenol derivate. After viable S. aureus was added, phagocytosis was stopped at different time points. Leukocytes were then stained with monoclonal antibodies for flow cytometric analysis of granulocyte recruitment (ratio of ingesting granulocytes) and phagocytosis activity (fluorescence intensity of ingested bacteria). Both barbiturates inhibited granulocyte recruitment and phagocytosis activity in a dose-dependent manner, whereas propofol did not affect any of the investigated variables. At concentrations higher than 7.6 x 10(-3) M (for thiopentone, p < .008) and 1.1 x 10(-3) M (for methohexitone, p < .04), granulocyte recruitment and phagocytosis activity were significantly inhibited. The calculated inhibitory concentrations (IC50) of thiopentone for granulocyte recruitment and for phagocytosis activity were 1.3 x 10(-2) M and 1.1 x 10(-2) M, respectively. The corresponding values for methohexitone were 3.6 x 10(-3) M and 1.1 x 10(-3) M. CONCLUSIONS: Our in vitro model points at substantially different effects of barbiturates and propofol on phagocytosis of S. aureus, which is one of the most important pathogens in patients who need neuroprotective therapy. The inhibitory effects of both barbiturates demonstrate a strong dose-dependency, with more pronounced effects for methohexitone. Impairment of phagocytosis activity was more pronounced than granulocyte recruitment.

Effects of intraoperative blood salvage on leukocyte recruitment to the endothelium.

BACKGROUND: The contamination of salvaged wound blood with activated leukocytes has been suspected to play a role in leukocyte-mediated tissue injury by increased adhesion to the endothelium. To verify this hypothesis, the authors performed a clinical study to examine the effects of blood salvage on leukocyte-endothelial interactions. METHODS: Expression of L-selectin, CD18, and CD11b and leukocyte adhesion to activated endothelium from human umbilical veins were measured in 25 patients undergoing major orthopedic surgery. Adhesion of fluorescently labeled leukocytes was examined in a flow chamber at shear rates of 50-1,600 s. Comparisons were made between samples from venous blood and from processed salvaged wound blood (SWB). RESULTS: At 30% hematocrit, SWB contained 2,162 +/- 147 leukocytes/microl. In comparison with venous blood, CD11b was up-regulated in SWB 1.3- to 3.6-fold on monocytes and neutrophils, whereas L-selectin and CD18 decreased on monocytes by 53% and 15%, respectively (P < 0.05). Despite up-regulation of CD11b, firm adhesion was significantly reduced by 74-76% in SWB. Rolling fractions and rolling velocities were significantly higher in SWB, and their relation to shear rate was markedly altered (P < 0.01). In addition, adherent leukocytes from SWB were significantly less resistant to increments of shear rate than leukocytes from venous blood (P < 0.01). CONCLUSIONS: Despite up-regulated CD11b, integrin-mediated adhesion is markedly impaired in salvaged blood. Therefore, the effect of blood salvage cannot be predicted from cell surface expression but rather from functional assays. The former hypothesis, that leukocytes from SWB aggravate leukocyte-mediated tissue injury by increased adhesion, may not be as great a concern as previously suggested.

Synthetic colloids attenuate leukocyte-endothelial interactions by inhibition of integrin function.

BACKGROUND: It has been suspected that synthetic colloids may interfere with leukocyte adhesion by down-regulation of endothelial cell adhesion molecules. Although inhibition of endothelial inflammation might reduce leukocyte-related tissue injury, the same mechanism may be detrimental for host defense during severe infection. Regarding the widespread use of colloids, the authors performed a laboratory investigation to determine the mechanisms by which synthetic colloids interfere with leukocyte-endothelial interactions. METHODS: Adhesion molecule expression on native and cytokine-activated endothelium from umbilical veins was measured after pretreatment with gelatin and various preparations of dextran or hydroxyethyl starch. Inhibition of neutrophil adhesion to activated endothelium was examined in a flow chamber by perfusion of untreated and colloid-treated neutrophils over colloid-pretreated endothelium at 2 dyn/cm. Comparisons were made between untreated controls, colloid-pretreated endothelium, and colloid-cotreated neutrophils. RESULTS: Intercellular adhesion molecule 1, vascular cell adhesion molecule 1, E-selectin, and P-selectin were not attenuated by any colloid. Accordingly, colloid pretreatment of endothelium alone did not reduce neutrophil adhesion. In contrast, when neutrophils were cotreated by addition of colloids to the perfusate immediately before perfusion, adhesion decreased by 31-51% (P < 0.05) regardless of the colloid type. As indicated by the twofold increased rolling fractions, this reduction was due to an inhibition of neutrophil integrins. CONCLUSIONS: This study shows that synthetic colloids inhibit neutrophil adhesion by a neutrophil-dependent mechanism rather than interfering with endothelial cell activation. This suggests that inhibition of leukocyte sequestration by volume support is a common and transient phenomenon depending on the colloid concentration in plasma.

The effects of fresh frozen plasma on neutrophil-endothelial interactions.

UNLABELLED: Leukocyte adhesion to endothelial cells contributes to microcirculatory disturbances during severe shock syndromes. Whereas certain plasma expanders inhibit leukocyte adhesion, contaminants of plasma protein solutions upregulate endothelial cell adhesion molecules in certain cases. We performed this study to determine whether fresh frozen plasma (FFP) affects neutrophil-endothelial interactions in cocultures of neutrophils and human umbilical vein endothelial cells (HUVEC) in vitro. HUVEC (n = 9) were incubated with either 20% FFP or 20% serum in medium for 6 h. Expression of E-selectin, intercellular adhesion molecule 1, and vascular cell adhesion molecule-1 was induced by tumor necrosis factor alpha (0.5 ng/mL for 4 h) and measured by flow cytometry. Neutrophil adhesion was examined in a parallel plate flow chamber in which isolated neutrophils were perfused over pretreated HUVEC under postcapillary flow conditions. Incubation with FFP decreased E-selectin and intercellular adhesion molecule 1 on activated HUVEC by 28% and 22%, respectively (P < or = 0.01; analysis of covariance). Consequently, neutrophil adhesion decreased by 20%-41% in FFP-treated cocultures (n = 4; P < or = 0.01; paired Student's t-test). We conclude that FFP attenuates the inflammatory response of endothelial cells with regard to neutrophil-endothelial interactions. Because the composition of patients' plasma is affected not only by transfusion, but more frequently by shock treatment with IV fluids, plasma dilution in critically ill patients could be important. IMPLICATIONS: During shock, fluid administration leads to a massive dilution of plasma. Apart from maintaining hemodynamics, this might affect tissue damage by influencing leukocyte accumulation in the microvasculature. Using endothelial cells, isolated neutrophils, and a parallel plate flow chamber, we studied the effects of fresh frozen plasma on neutrophil-endothelial interactions.

Local anesthetics impair human granulocyte phagocytosis activity, oxidative burst, and CD11b expression in response to Staphylococcus aureus.

BACKGROUND: With invasion of bacteria, the host defense system is activated by a complex cascade of various mechanisms. Local anesthetics previously were shown to interact with diverse components of the immune response, such as leukocyte adherence on endothelial monolayers, oxidative burst, or crosstalk within lymphocyte subset populations. However, effects of newer local anesthetics like bupivacaine and ropivacaine on antibacterial host defense-primarily phagocytosis activity, oxidative burst, or CD11b expression-still remain unclear. METHODS: Whole blood samples were preincubated with local anesthetics (lidocaine, 9.2, 92.2, and 1,846 microm bupivacaine, 6.1, 61, and 770 microm; ropivacaine, 6.4, 64, and 801 microm). For the oxidative burst and CD11b assay, dihydroethidium was added to the probes. After viable Staphylococcus aureus was added in a 5 to 1 ratio following leukocyte count, phagocytosis was stopped at different times, and staining with monoclonal antibodies was performed for subsequent flow cytometric analysis of phagocytosis activity, oxidative burst, and CD11b expression. RESULTS: Granulocyte phagocytosis activity, CD11b expression, and generation of reactive oxygen species were significantly reduced by lidocaine (P < 0.0002) and bupivacaine (P < 0.005) in the highest concentration (1,846 microm and 770 microm, respectively). The capability of granulocytes to ingest bacteria was significantly depressed only by lidocaine (P < 0.003). Ropivacaine had no significant effect on any parameter investigated. CONCLUSIONS: Local anesthetic dose and structure dependently inhibit inflammatory and immunologic parameters of granulocyte functions. Ropivacaine shows low interference with granulocyte immunologic and inflammatory functions.