Effects of zymosan-activated plasma and phorbol myristate acetate on isolated, perfused rabbit lungs

The effects of complement activation on pulmonary vascular permeability are disputed. In rabbit lungs perfused with autologous blood, zymosan activated plasma (ZAP) induced a moderate increase in pulmonary vascular resistance (PVR), but did not detectably change the vascular permeability within 2 h. The stronger neutrophil granulocyte (PMN) activator, phorbol myristate acetate (PMA), usually gave larger PVR increases and also increased pulmonary vascular permeability. Lungs from neutropenic animals, similarly perfused and given PMA, showed unchanged PVR reactions but had no apparent increase in vascular permeability. Lungs perfused with cell-free medium and given PMA displayed modest PVR increases, and no measurable permeability change. The lung preparatory procedure itself markedly influenced leukocyte circulation. Exsanguination of lung donors decreased the concentration of circulating PMN significantly, and they virtually disappeared from the perfusate within minutes after start of lung perfusion. PMN-mediated effects must therefore have been caused by cells already sequestered in the lungs. We conclude that ZAP does not induce an increased pulmonary vascular permeability in isolated, perfused rabbit lungs, in contrast to PMA. The permeability effects of PMA appear to be PMN dependent.     

Complement Activation and its Relationship to Adult Respiratory Distress Syndrome. An Experimental Study in Pigs

Pulmonary leucostasis induced by complement activation has been considered an important pathogenic factor in adult respiratory distress syndrome (ARDS). To determine whether complement activation per se could evoke pulmonary dysfunction similar to ARDS, pigs were repeatedly infused with complement-activated plasma (CAP). Complement activation was produced by incubation of plasma with zymosan. Three groups of animals were investigated. Control animals received non-activated plasma. Nine animals (Group II) were given four infusions of GAP at a rate of 7 ml min^-1, and another nine animals (Group III) received two CAP infusions at a rate of 7 ml min^-1 followed by two at a rate of 14 ml min^-1. In the control animals there were no changes in gas exchange or haemodynamic variables and the leucocyte counts gradually increased. Infusion of CAP resulted in transient peripheral leucopenia and a dose-rate-dependent reversible increase in pulmonary vascular resistance in all animals. In one animal of Group II and in six of Group III there was a significant infusion-related decrease in PaO₂ due to increased venous admixture. These animals were characterized by an enhanced pulmonary vascular tone before the start of the first CAP infusion. They also displayed a more pronounced pulmonary vascular response to infusion of CAP. The changes in gas exchange variables and pulmonary haemodynamics showed no relation to the degree of leucopenia or decrease in platelet count. The increased venous admixture was caused by "dry" ventilation/perfusion mismatching and not by oedema. These results suggest that additional factors besides complement activation and pulmonary leucostasis are required for the development of increased microvascular permeability and the pulmonary oedema characterizing ARDS.     

Neutrophil activation in sepsis. The relationship between fmet-leu-phe receptor mobilization and oxidative activity

To elucidate further the manifestations and mechanisms of neutrophil (PMN) activation, PMNs from control and septic subjects were studied at baseline and under conditions of graded, in vitro activation. At baseline (4 degrees C PMN isolation), septic-derived PMNs were activated, as manifested by twofold increases in fmet-leu-phe (FMLP)-induced oxidative activity and concomitant FMLP surface receptor expression, compared with controls. Following degranulationlike maximal activation (phorbol myristate acetate pretreatment), both PMN populations exhibited maximal FMLP-induced oxidative priming and receptor up-regulation. However, following exudation-like moderate activation (37 degrees C pretreatment), control PMNs underwent significant receptor mobilization and oxidative priming but septic-derived PMNs exhibited oxidative deactivation (decreased FMLP-induced oxidative activity) without changes in FMLP receptor expression. Our data support the theory that while circulating PMNs in sepsis may promote oxidant-related microvascular lung injury, their oxidative deactivation following transpulmonary exudation (simulated by 37 degrees C pretreatment) may underlie the increased incidence of pulmonary infections seen in sepsis-induced adult respiratory distress syndrome.     

A Porcine Model of Early Adult Respiratory Distress Syndrome Induced by Endotoxaemia

To study the pathophysiology of early adult respiratory distress syndrome (ARDS) induced by sepsis, spontaneously breathing pigs under ketamine anaesthesia were investigated. Twenty animals were infused i.v. with E. coli endotoxin (10 micrograms . h-1 . kg-1) over 6 h, and ten control animals received physiological saline. In the controls, cardiac output (Qt) and O2 delivery decreased slightly. There were no changes in pulmonary gas exchange, pulmonary haemodynamics or extravascular lung water (EVLW). The polymorphonuclear (PMN) leucocyte count gradually increased, while the platelet count decreased slightly. Endotoxin infusion caused profound deterioration of pulmonary gas exchange, a marked rise in pulmonary vascular resistance (PVR) and a moderate increase in EVLW. The pulmonary dysfunction was not attributable to the pulmonary oedema per se, whereas a "dry" ventilation/perfusion inequality played an important role. The "responders" (peak venous admixture greater than 20%; n = 14) were characterized by higher Qt and lower PVR than the "non-responders". Qt declined progressively, especially in non-survivors. O2 delivery decreased considerably. Metabolic acidosis probably indicated oxygen deficit. Eleven of 20 animals died during the observation period. Mortality was related more to the imbalance between O2 delivery and oxygen demand than to the deterioration in pulmonary gas exchange. The PMN count decreased markedly while the gradual decline in platelet count was similar to that in the controls. Lung microscopy revealed PMN accumulation in the microvasculature, moderate interstitial oedema and microvascular blood stasis. Our porcine model, which closely mimics early ARDS in man, will be useful in further studies of the pathophysiological pathways and the treatment of this syndrome.     

Alterations of polymorphonuclear leukocyte glycogen metabolism and glucose uptake in dialysis patients

Human polymorphonuclear leukocytes (PMNs) are activated during extracorporeal circulation. An indicator of PMN activation may be the glycogen-degrading enzyme phosphorylase. It is unknown whether dialysis therapy may influence PMN carbohydrate metabolism. Therefore, PMNs were isolated from healthy control subjects, patients undergoing continuous ambulatory peritoneal dialysis (CAPD), and patients undergoing regular hemodialysis therapy (RDT) before, during, and at the end of hemodialysis (HD) treatment using dialyzers made of polysulfone or polymethylmethacrylate (PMMA). Nifedipine (NIF) was continuously infused during HD with PMMA in 5 patients at a dose of 18 micrograms/kg body weight per hour. Glycogen, activity of glycogen synthetase and phosphorylase (active and inactive forms of both enzymes), and glucose uptake with and without stimulation with the chemotactic peptide FMLP were determined in these PMNs. During HD with PMMA, there was a significant increase of PMN phosphorylase "a" activity 15 and 30 minutes after the start of HD. HD with polysulfone did not stimulate the active "a" form of the glycogen-degrading enzyme in PMNs. HD with PMMA significantly inhibited the active I-form of glycogen synthetase, whereas polysulfone activated glycogen synthetase I. NIF inhibited phosphorylase "a" activation during HD with PMMA. PMN glycogen content and glucose uptake were improved during HD with polysulfone, but not with PMMA. PMN glycogen content, activity of glycogen synthetase, and glucose uptake were significantly lower also in CAPD patients compared with healthy controls. These data show that HD with PMMA activates PMN glycogenolysis. This effect can be inhibited by calcium channel blockers. PMN glycogen content of RDT and CAPD patients is significantly lower compared with healthy controls due to inhibition of glycogen synthesis. Elimination of dialyzable factor(s) improves, but does not restore, PMN glycogen synthesis and glucose uptake.     

Plasma from aged stored red blood cells delays neutrophil apoptosis and primes for cytotoxicity: abrogation by poststorage washing but not prestorage leukoreduction

BACKGROUND: Blood transfusion-particularly that of older stored red blood cells (RBCs)--is an independent risk factor for postinjury multiple organ failure. Immunomodulatory effects of RBC transfusion include neutrophil (PMN) priming for cytotoxicity, an effect exacerbated by longer RBC storage times. We have found that delayed PMN apoptosis in trauma patients is provoked by transfusion, independent of injury severity. We hypothesized that aged stored RBCs delay PMN apoptosis, but that prestorage leukodepletion or poststorage washing could abrogate the effect. METHODS: Healthy volunteers each donated 1 unit of blood. One half was leukodepleted, and RBC units were processed in the usual fashion and stored at 4 degrees C. Aliquots were removed on days 1, 14, 21, and 42 and the plasma fraction isolated. Selected aliquots were washed with normal saline before plasma isolation. PMNs harvested from healthy controls were incubated (5% CO2, 37 degrees C) with unmodified, leukoreduced, or washed RBC plasma (20% plasma/80% RPMI 1640), and apoptosis assessed by morphology after 24 hours. Apoptotic index (apoptotic PMNs/total PMNs) was compared. PMN priming for superoxide release was also assessed after plasma exposure. RESULTS: PMN apoptosis was delayed by RBCs stored for 21 or 42 days. Prestorage leukodepletion did not alter the effect. However, washing 42-day-old RBCs abrogated the effect. PMN priming for superoxide was provoked by stored packed RBCs in an identical pattern to delayed apoptosis. CONCLUSION: Plasma from stored RBCs-even if leukoreduced-delays apoptosis and primes PMNs. The effect becomes evident at 21 days and worsens through product outdate (42 days), but may be prevented by poststorage washing. Inflammatory agents contaminating stored blood likely mediate the effect. Modification of transfusion practices (e.g., giving fresher or washed RBCs or blood substitutes) may attenuate adverse immunomodulatory effects of transfusion in trauma patients.     

Clinically relevant hypertonicity prevents stored blood- and lipid-mediated delayed neutrophil apoptosis independent of p38 MAPK or caspase-3 activation

BACKGROUND: Delayed apoptosis of primed neutrophils (PMNs) may facilitate PMN-mediated tissue injury leading to postinjury multiple organ failure. Aged (42-day-old) stored red blood cells (RBC42) delay PMN apoptosis through proinflammatory phospholipids such as platelet-activating factor (PAF) and lyso-phosphatidylcholine (LPC). Hypertonic saline (HTS) attenuates PMN cytotoxic functions. We hypothesized that clinically relevant HTS would provoke PMN apoptosis, as well as prevent stored blood- and lipid-mediated delayed PMN apoptosis through activation of p38 mitogen-activated protein kinase (MAPK) and caspase-3. METHODS: PMNs harvested from healthy volunteers were incubated (5% CO(2), 37 degrees C, 24 hr) with RBC42 plasma, PAF (20 microM), or LPC (4.5 microM), with or without the p38 MAPK inhibitor SB 203580, the caspase-3 inhibitor zDEVD-fmk (10 micromol/L) or the pan-caspase inhibitor zVAD-fmk (20 micromol/L). Duplicate samples were preincubated in HTS (Na [180 mM]). Apoptotic index (% PMNs undergoing apoptosis) was assessed morphologically. p38 MAPK activation was assessed by Western blotting. Caspase-3 activity was measured colorimetrically. RESULTS: PAF, LPC, and RBC42 plasma delayed apoptosis; HTS increased apoptosis compared with controls. HTS prevented PAF, LPC, and RBC42-delayed apoptosis. p38 MAPK was not activated by HTS; its inhibition had no effect on the actions of HTS. Caspase inhibition attenuated the ability of HTS to increase apoptosis, but it did not affect the ability of HTS to restore healthy PMN apoptosis in the presence of RBC42. CONCLUSION: HTS increases PMN apoptosis and prevents stored blood- and lipid-mediated delayed PMN apoptosis. HTS may activate caspase-3, but alternative signaling pathways appear to be involved in modulating the effects of lipids on PMN apoptosis.     

Granulocytes in bronchial lavage fluid after major vascular surgery

Increased numbers of polymorphonuclear granulocytes (PMN) in the airways, as measured by PMN content in bronchial lavage fluid (P less than 0.01), were found 3 h postoperatively in ten patients undergoing surgery for lumbar aortic aneurysms. An increase in plasma levels of the complement split product C3dg from 6 (0-19) AU/ml preoperatively to 20 (13-50) AU/ml 3 h after surgery (P less than 0.01), indicates an activation of the complement cascade. These changes were not accompanied by increased elastase activity in the bronchial lavage fluid or by major changes in pulmonary blood gas exchange or vascular resistance, indicating that massive PMN activation, analogous to that proposed in adult respiratory distress syndrome (ARDS) had not taken place. In conclusion, complement system activation and migration of PMN into the airways, as seen in connection with major vascular surgery, does not seem to contribute to ARDS-type pulmonary dysfunction.     

术中保温对肺癌根治术患者中性粒细胞呼吸爆发的影响

Objective To investigate the influence of intraoperative thermostasis over respiratory burst of polymorphonuclear neutrophils (PMNs) in patients undergoing radical operation for lung cancer.Methods Thirty-two ASA Ⅱ or Ⅲ patients scheduled for radical operation for lung cancer under general anesthesia were randomized into two groups ( n = 16 each): control group (Group C) and warming group (Group W). The patients in Group C were kept warm by routine measures such as using woollen blankets, while the patients in Group W were kept warm by force-air warming system and fluid warming device as soon as the patients were admitted to the operation room. Rectal and axillary temperatures were continuously monitored as the core and surface temperature, respectively. The core temperature was maintained at the preoperative level (baseline). Anesthesia was induced with midazolam, fentanyl and propofol. Tracheal intubation was facilitated with rocuronium. Anesthesia was maintained with isoflurane and nitrous oxide and intermittent i.v. boluses of fentanyl, midazolam and vecuronium. Venous blood samples were obtained before, during and at the end of surgery for normal blood analysis and respiratory burst of PMNs which included activated PMNs count and reactive oxygen species (ROS) production.Results (1) WBC and PMN counts were significantly increased during and after operation as compared with the baseline values before operation in both groups and there was no significant difference in WBC and PMN counts between the two groups. (2)Phorbol-12-myristate-13-acetate (PMA) stimulation resulted in higher intraoperative and postoperative activated PMN counts in both groups and higher postoperative ROS production in Group W. Postoperative ROS production was significantly higher in Group W than in Group C. (3) The PMN counts without stimulation activation during operation and intra- and post-operative ROS production were significantly decreased as compared with the baseline values before operation in Group C, while in Group W there was no significant difference in pre-, intra- and post-operative activated PMN counts and ROS production. The intraoperative PMN counts and intra- and post-operative ROS productions were significantly higher in Group W than in Group C.Conclusion Intraoperative thermostasis can effectively maintain activated PMN count and ROS production without stimulation and enhance ROS production with stimulation in patients undergoing radical operation for lung cancer.     

Effect of local and systemic burn microenvironment on neutrophil activation as assessed by complement receptor expression and morphology

Previously, we documented that humoral factors, especially complement products, contained in burn wound blister fluid (BF) modulate normal neutrophil (PMN) function and metabolism. The goal of the current study was to examine the effects of the local (BF) and systemic (burn serum or plasma) burn microenvironment on PMN activation as assessed by complement receptor expression and morphology. Induction of CR1 (C3b) and CR3 (iC3b) receptor expression of normal PMNs after incubation in medium, BF, or plasma (serum) from healthy volunteers or burned patients was measured using monoclonal antibodies and flow cytometry. BF (10% v/v, 37 degrees C) induced 25% more CR1 receptor expression than control or burn plasma (p less than 0.05), while the levels of CR3 expression of PMNs incubated in BF (10% or 80% v/v at 37 degrees C) was more than 300% that found when the PMNs were incubated in medium, burn, or patient plasma (p less than 0.05). The electron microscopic appearance of PMNs incubated in these fluids documented that degranulation was greater when cells were incubated in BF than medium, control, or patient plasma. These results indicate that PMN activation (CR1 and CR3 expression) is greater in cells exposed to the local (BF) than the systemic (plasma) humoral microenvironment shortly after thermal injury.     

CD18 adhesion receptors, tumor necrosis factor, and neutropenia during septic lung injury.

Sequestration of neutrophils (PMNs) in the pulmonary microvasculature and associated neutropenia are characteristic features of experimental models of septic lung injury. The etiology of altered PMN kinetics during septic lung injury is uncertain, but may be partially due to increased adhesiveness of activated PMNs to pulmonary endothelium. This study examines the relationship between the expression of PMN CD18 adhesion receptors, the evolving neutropenia, and plasma tumor necrosis factor (TNF) activity in a porcine model of septic lung injury. Acute lung injury was induced by infusion of live Pseudomonas aeruginosa (5 x 10(8) CFU/ml at 0.3 ml/20 kg/min) for 60 min (Group Ps, n = 6). Control animals (Group C, n = 3) received a 60-min infusion of sterile 0.9% saline. CD18 expression of circulating PMNs was measured by quantitative immunofluorescent flow cytometry. Plasma TNF activity was measured by L929 fibroblast cytolytic assay. Group Ps developed a significant neutropenia by 30 min (14.9 +/- 2.5 vs 23.4 +/- 3.3 x 10(3) cells/microliter at baseline, P less than 0.05, ANOVA) with circulating neutrophils exhibiting significantly increased CD18 expression by 60 min (6.34 +/- 0.72 vs 5.01 +/- 0.52 equivalent soluble fluorescence molecules (ESFM) x 10(3) at baseline, P less than 0.05, ANOVA). Group Ps demonstrated a significant increase in plasma TNF activity by 30 min (2.5 +/- 0.9 vs 0.7 +/- 0.3 U/ml at baseline). There was no significant change in PMN count, PMN CD18 expression, or plasma TNF activity in Group C. In complimentary in vitro studies, porcine PMNs stimulated with recombinant human TNF-alpha (n = 5) demonstrated a time- and dose-dependent increase in CD18 expression.(ABSTRACT TRUNCATED AT 250 WORDS)     

Polymorphonuclear leukocyte rigidity is defective in patients with chronic renal failure.

BACKGROUND: The purpose of the study was to investigate the rigidity of polymorphonuclear leukocytes (PMNs) in non-dialysed chronic renal failure (CRF) and haemodialysis (HD) patients. METHODS: PMN rigidity as well as tumour necrosis factor alpha (TNF-alpha) and interleukin 1beta (IL-1beta) plasma levels were assessed in 10 early-stage CRF, 10 late-stage non-HD, and 10 HD patients, before and during dialysis. In HD patients both cellulose acetate and polysulphone membranes were used. Ten healthy subjects served as controls. Rigidity was tested by counting the deformability in morphologically passive PMNs by the micropipette method. Cytokine levels were measured by enzyme-linked immunosorbent assay. RESULTS: PMN rigidity was significantly increased in end-stage CRF patients regardless of HD but not in early-stage CRF. In HD patients PMN rigidity increased significantly 60 min after initiation of HD. There was an increase of TNF-alpha and IL-1beta levels in end-stage non-HD and HD patients and a further increase at 60 min after initiation of HD. The percentage of morphologically activated PMNs was increased only during dialysis. The nature of the HD membrane had no influence on rigidity, PMN activation, or cytokine production. CONCLUSIONS: The results indicate that PMN rigidity is defective in end-stage chronic CRF patients and is further increased 60 min after initiation of HD, regardless of the nature of the HD membrane used. PMN activation, increased TNF-alpha and IL-1beta levels, or a direct PMN impairment may cause the observed cell rigidity.     

Effects of methylprednisolone on complement activation and leukocyte counts during cardiopulmonary bypass

Generation of the complement activation products C3dg and terminal complement complex (TCC) and numerical changes in peripheral granulocytes (PMN) and lymphocytes were assessed in patients undergoing aortocoronary bypass surgery with extracorporeal circulation (ECC). Fluid from bronchial lavage performed preoperatively and 4 hours postoperatively was analyzed for granulocyte elastase activity and PMN content. Ten of the 20 patients received methylprednisolone (30 mg/kg b.w.) immediately before ECC. No difference was found between them and the control group regarding C3dg and TCC, and both groups showed similar postoperative decrease of peripheral blood lymphocytes. The postoperative PMN count in peripheral blood was significantly higher in the methylprednisolone group than in the controls from 12 hours onwards. In bronchial lavage fluid the postoperative PMN count was unaltered in the methylprednisolone group, but significantly increased in the controls. No granulocyte elastase activity was found before or after surgery in either group. The results indicated that methylprednisolone does not affect complement activation during cardiopulmonary bypass, but increases the granulocytes in peripheral blood postoperatively.     

Extracellular signal-related kinase 1/2 and p38 mitogen-activated protein kinase pathways serve opposite roles in neutrophil cytotoxicity.

BACKGROUND: Inflammatory stimuli rapidly activate mitogen-activated protein kinases (MAPKs) in neutrophils (PMNs). However, their role in cytotoxic function remains unknown. Elucidating the signals involved in release of cytotoxic agents from PMNs may provide new avenues for therapy in diseases of diminished or excessive PMN function. HYPOTHESIS: The p38 MAPK and extracellular signal-related kinase 1/2 (ERK1/2) modulate superoxide generation and elastase release in activated human PMNs. STUDY DESIGN: Isolated human PMNs were incubated with specific inhibitors of MAPK pathways, or vehicle control solution, before activation with the bacterial peptide f-Met-Leu-Phe. MAIN OUTCOME MEASURES: The rate of superoxide release from activated PMNs was measured by the superoxide dismutase-inhibitable reduction of cytochrome-c. Elastase release from PMNs was determined by cleavage of the substrate Ala-Ala-Pro-Val-pNA. RESULTS: Superoxide release from activated PMNs was inhibited by blockade of p38 MAPK activation but unaffected by blockade of ERK1/2. Conversely, elastase release was unaffected by p38 MAPK inhibition and increased by ERK1/2 inhibition. CONCLUSIONS: Activation of p38 MAPK promotes superoxide release from PMNs activated by f-Met-Leu-Phe. The ERK1/2 pathway may serve as a negative feedback mechanism for granule exocytosis.     

Effects of high dose E. coli lipopolysaccharide on rabbit lung function,and of subsequent addition of chemotactic granulocyte activators to their isolated,blood-perfused lungs

E. coli LPS was infused (1 μg/kg to 5 mg/kg over 30 min) to spontaneously breathing rabbits, and their arterial blood pressure (ABP), blood leukocyte count and blood gases were observed for 2.5–3.5 h. Pulmonary vascular and airway function were subsequently evaluated in vitro by comparing weight changes, fluid filtration rates, pulmonary vascular resistance (PVR) and airway pressures in their isolated, blood-perfused lungs with those in lungs from untreated rabbits. Lung preparations from both groups of animals were then exposed to autologous zymosan-activated plasma (ZAP) or n-formyl-methionyl-leucyl-phenylalanine (FMLP) and perfused for 2 more hours. LPS addition to isolated rabbit leukocytes increased cell aggregation; cell chemiluminescence after activation with FMLP was also enhanced. Infusion of 1–5 mg/kg LPS decreased the count of all types of leukocytes and caused a metabolic acidosis (BE< –8 mmol), but no decrease in ABP. PAo₂-Pao₂ increased by about 2.0 kPa. No vascular permeability increase was detected in the lungs of these animals during subsequent in vitro perfusion. Addition of ZAP or FMLP during perfusion markedly increased the PVR in lungs from LPS animals, but did not induce major microvascular leakage. No significant differences in edema between lungs from LPS-treated and control animals were found by microscopy.     

Attenuation of intra-operative surgical stress response has no influence on post-operative degranulation of polymorphonuclear granulocytes.

Degranulation of polymorphonuclear (PMN) granulocytes, complement activation, and the endocrine metabolic response to hysterectomy were compared in two groups of patients receiving either general anaesthesia (GA group) or combined epidural analgesia and general anaesthesia (GAE group). The B-leucocyte and the cortisol responses were attenuated in the GAE group. There was no sign of complement activation. The post-operative rise in elastase-alpha-1-proteinase levels 4 h post-operatively on the first, second and third post-operative days was similar in the two groups. Plasma lactoferrin values were significantly elevated 4 h post-operatively and reached pre-operative values on Day 1 in both groups. Attenuation of the surgical stress response during uncomplicated hysterectomy did not influence the post-operative increase in proteinase activity. Mediators generated at the site of surgical trauma may account for the PMN degranulation observed after major surgery.     

Stimulation of neutrophil activation during coronary artery bypass grafting: comparison of crystalloid and blood cardioplegia

BACKGROUND: During myocardial ischemia, activation of polymorphonuclear neutrophils (PMNs) results in the production of free oxygen radicals, which increase myocardial injury. It has been shown that PMNs also produce nitric oxide. It is not clear whether PMNs become activated as a result of their direct contact with ischemic/reperfused myocardium or if PMN activation and free oxygen radical production are effects of specific stimuli released during coronary artery bypass grafting (CABG). The aim of the current study was to evaluate plasma-mediated neutrophil stimulation and production of superoxide anion (O2) and nitric oxide in patients undergoing CABG, and to verify whether crystalloid and blood cardioplegia can modify such stimulation. METHODS: Coronary sinus, peripheral arterial, and venous plasma samples were collected from 50 patients who underwent CABG and were divided into 2 equal groups which received either crystalloid or blood cardioplegia: directly before myocardial ischemia and aortic cross-clamping; at the beginning of reperfusion after aortic clamp release; and 30 minutes after reperfusion. O2 and nitric oxide production by PMN was evaluated by standard methods. RESULTS: There was a significant (p < 0.05) increase in O2 production by PMN incubated with plasma obtained from the coronary sinus immediately after reperfusion in patients receiving crystalloid cardioplegia compared to blood cardioplegia. No difference was observed in plasma stimulation of nitric oxide production by PMN in the 2 groups of patients at different times during the procedure. CONCLUSIONS: Cardioplegia may affect release of neutrophil-oriented stimuli from ischemic myocardium and modify neutrophil activation during coronary artery bypass grafting.     

Hypoxaemia created by pulmonary oedema after pulmonary microemboli in dogs

Pulmonary microemboli may play a role in the adult respiratory distress syndrome, creating both pulmonary oedema and hypoxaemia. We measured the time course of pulmonary oedema and hypoxaemia after pulmonary microemboli of 63-74 mu starch were infused into dogs. Dogs were divided into two groups: six dogs that did not develop pulmonary oedema, and seven that did. Immediately after emboli there was no difference between groups in the large fall in PaO2 or the rise in Qs/Qt. Therefore the hypoxaemia of pulmonary embolism is not created by pulmonary oedema. As pulmonary oedema increased with time in the oedematous dogs, PaO2 fell further. There was no further reduction in PaO2 in the dogs who did not develop pulmonary oedema. We conclude that hypoxaemia after pulmonary embolism may be worsened if pulmonary oedema occurs, but the immediate large reduction in PaO2 after embolism is not created by pulmonary oedema.     

Neutrophil activation after burn injury: contributions of the classic complement pathway and of endotoxin

We attempt to elucidate the mechanisms of neutrophil (PMN) activation after burn injury. We previously reported prolonged elevations of PMN cell surface complement (C) opsonin receptor levels after burn trauma with a corresponding period of depressed PMN chemotaxis to C5a, which suggests that the C product, C5a, was responsible for PMN activation. However, a lack of direct correlation of C activation with C receptor levels soon after injury raised the possibility of a second PMN-activating substance. We therefore investigated the effect of endotoxin (LPS) on the expression of the C receptors (CR1 and CR3) by normal human PMNs. Concentrations from 0 to 50 ng/ml of LPS 026:B6 caused a dose response increase in the PMN surface expression of CR1 and CR3 as assessed by monoclonal antibody binding and indirect immunofluorescence. The relative CR1-dependent fluorescence rose from a mean of 50 to 385 and CR3 from 50 to 300. Chelation by ethylenediaminetetra acetic acid (EDTA) did not influence this dose response, thus ruling out the possibility of C activation by LPS--an inference supported by the lack of complement activation observed with these concentrations of LPS in normal serum. A similar dose response was obtained in the absence of other cell types or serum, which implies a direct effect that mimicked that of C5a. To determine the mechanism of the later, prolonged C activation after burn injury, we next examined C activation products in 22 patients with burn injuries. Elevations of plasma C3a desArg were present and persisted for 50 days. Elevations were at maximum levels on days 9 through 13 postburn (mean +/- standard error of mean [SEM], 496 +/- 47 ng/ml versus normal 113 +/- 32; p less than 0.01). These were accompanied by elevations of C4a desArg (917 +/- 154 ng/ml versus normal 424 +/- 50; p less than 0.01), which are indicative of classic pathway activation. Finally, we examined PMN function, phagocytosis and percentage killing of Staphylococcus aureus, and found PMN function to be unaltered in the 22 patients. Thus PMN activation after burn injury appears to be caused by LPS soon after injury and by C5a later after injury and affects only selected PMN functions.     

The pathogenesis of pulmonary contusion: an open chest model in the rat

BACKGROUND: Chemokines direct leukocytes to areas of inflammation or injury. In general, CC chemokines (MCP-1, MIP-1alpha, RANTES) are chemoattractants for mononuclear cells and CXC (CINC-1, MIP-2alpha) for polymorphonuclear cells (PMNs). Herein we describe an open chest model of pulmonary contusion (PC) in a rodent (rat) and have identified a possible role for CC and CXC chemokines in the pathogenesis of PC. METHODS: Sprague-Dawley rats (350 g) underwent thoracotomy. The exposed lung was struck with a piston at 5.2 m/s (150 J/M2). Blood, bronchoalveolar lavage (BAL), and lung tissue were collected at 3 hours and 24 hours after injury. PaO2/FiO2 (P/F) ratio was calculated at 15-minute intervals for 3 hours after contusion. Serum was evaluated for cytokine and chemokine expression using ELISA. Cell count/differential was performed on BAL, and lung tissue was obtained for histologic analysis, protein expression and wet to dry weights. Data are reported as pg/mL +/- SE. Data were analyzed using Student's t test to identify significant differences (p < or = 0.05 significant) between sham and injured animals. RESULTS: Piston impact caused PC based upon morphologic and histologic criteria. BAL cell count and lung wet to dry weights were increased and P/F ratio was decreased after PC. Systemic levels of IL-ra, MCP-1, and the CXC chemokines MIP-2alpha and CINC-1 were significantly elevated at 3 hours when sham and injured animals were compared. All chemokines were found to be significantly elevated at 24 hours, consistent with the early PMN and subsequent mononuclear infiltration observed in the contused lung. Pulmonary expression of TNF-alpha, IL-1beta, CINC-1, MIP-2alpha, ICAM-1, and elastase were increased and activated systemic neutrophils showed increased CD-11b. CONCLUSION: A model of PC is described in which innate inflammation is activated locally and systemically. Systemic levels of CC and CXC chemokines are increased after PC. This correlates with elevated PMN CD-11b expression, enhanced pulmonary ICAM-1 expression, and mononuclear and PMN infiltration into the lung and alveolar space. Elevated levels of CC and CXC chemokines are seen after PC and may be involved in the lung's inflammatory response to injury.