Elemental diet alters macrophage function in mice

Administration of a chemically defined liquid elemental diet (ED) induces spontaneous bacterial translocation to mesenteric lymph nodes (MLN) in animal models. The influence of this process on host immunity is unclear. This study evaluated the effects of ED on peritoneal macrophage (PM phi) antimicrobial functions. Conventional C57/BL6 mice and endotoxin-resistant C3H/HeJ mice (n = 60) were randomized to be pair-fed either an ED or regular chow diet (RD) for 14 days. Blood, spleen, liver, and MLN were cultured for bacteria. PM phi were harvested for: percentage Candida albicans (CA) phagocytosis, percentage killing of CA, PM phi superoxide anion (O2-) production, and TNF-dependent macrophage cytotoxicity. Enteral feeding of ED in conventional C57/BL6 mice caused significant bacterial translocation to MLN but not other organs. Significant impairment of CA killing by PM phi occurred in the ED group and was associated with reduced O2- production. Tumor necrosis factor (TNF)-dependent cytotoxicity of PM phi was also decreased. In endotoxin-resistant C3H/HeJ mice, bacterial translocation was not observed and PM phi antifungal functions remained similar in both RD and ED groups. Thus, enteral feeding of an elemental diet downregulates host oxidative and antimicrobial mechanisms and TNF-dependent cytotoxicity in conventional mice which may be secondary to elemental diet-induced bacterial translocation.     

Effects of laparotomy on systemic macrophage function.

Surgical trauma induces immunosuppression that may adversely influence survival. This study examined the effect of laparotomy on two different macrophage populations, peritoneal macrophages (PM phi) and Kupffer cells. Female, 6- to 8-week old, CFW/C3H-HeN mice (n = 160) were randomly allocated to one of three study groups: control, ether anesthetic only, or ether anesthetic and laparotomy. On postoperative days 1 and 3, PM phis and Kupffer cells were harvested and assayed for superoxide anion production (O2-), percent macrophage phagocytosis of Candida albicans (CAP), percent C. albicans killed by macrophages (CAK), percent major histocompatibility complex (MHC)-class II antigen expression, and antigen presentation. Macrophages isolated on postoperative day 1 were also cocultured with 100 units/10(6) cells/ml interferon-gamma (IFN-gamma). Laparotomy significantly impaired microbicidal activity (O2-, percent CAP, and percent CAK) and antigen presentation on postoperative day 1. On postoperative day 3, O2- and antigen presentation were increased significantly (p less than 0.05) over control values, indicating a rebound phenomenon. Kupffer cell microbicidal function was unchanged on postoperative days 1 and 3. The initial immune impairment (PM phis: O2-, CAP, and CAK) was abrogated by IFN-gamma treatment. In immunosuppressed hosts after injury, administration of macrophage-activating factors such as IFN-gamma could be of therapeutic benefit.     

Impairment of pulmonary macrophage function with total parenteral nutrition.

OBJECTIVE: The effects of total parenteral nutrition (TPN) administration on pulmonary macrophage function and host response to gram-negative pulmonary infection were evaluated. SUMMARY BACKGROUND DATA: Administration of TPN resulted in increased infectious complications in traumatized and perioperative patients, but underlying mechanisms are unclear. METHODS: Twenty-six male Wistar rats underwent central vein cannulation and were randomized to isocaloric feeding of a regular chow diet (RD) plus saline infusion or TPN without chow diet for 7 days. Pulmonary alveolar macrophage (PAM phi) superoxide production, Candida albicans phagocytosis and killing, and tumor necrosis factor (TNF) production in response to endotoxin (LPS) were assessed. Mesenteric lymph nodes (MLN) were cultured. A second group of rats (n = 6/group) were inoculated intratracheally with a sublethal dose of 9 x 10(9) live Escherichia coli per animal, and the lungs were cultured quantitatively 72 hours later to assess bacterial clearance. Finally, 11 RD-fed rats and 13 TPN-fed rats received intratracheal inoculation of 1.4 x 10(10) live E. coli and were included in follow-up. RESULTS: Administration of TPN was associated with a significant increase in bacteria positive MLN compared with those in the RD group (p < 0.01). Pulmonary alveolar macrophage superoxide production, Candida albicans phagocytosis and killing, TNF production, and pulmonary clearance of bacteria were decreased significantly in TPN-fed rats compared with those fed a regular chow diet (p < 0.05). These pulmonary macrophage function changes were associated with a significantly higher mortality in TPN-fed rats compared with RD-fed rats after higher dose pulmonary E. coli inoculation. CONCLUSIONS: Defective host pulmonary antimicrobial immune responses during TPN are associated with intestinal bacterial translocation, and may explain increased infectious complications.     

Immunosuppressive mechanisms in protein-calorie malnutrition.

Protein-calorie malnutrition (PCM) induces immunosuppression leading to increased mortality rates. Impaired macrophage respiratory burst activity (superoxide anion [O2-] generation) occurs in PCM, but cellular mechanisms are unclear. The major pathway resulting in O2- production involves inositol lipid-dependent signal transduction. This study examined the effect of mild versus severe PCM on macrophage O2- generating signal transduction pathways specific for responses to Candida albicans. Mice (CFW/Swiss Webster: n = 300) were randomized to either control or low protein diets for 3 or 8 weeks. Peritoneal macrophages were harvested for O2- production, mannose-fucose receptor (MFR) expression, membrane phospholipid analysis, arachidonic acid (AA) content, prostaglandin E2 (PGE2) production, and protein kinase C levels. O2- release was impaired in both mild and severe PCM. MFR expression was also decreased at these time points. Inositol lipid content was significantly lower at the 8-week time point only, although PGE2 and AA were significantly higher in the low protein diet group at 3 weeks. Protein kinase C levels were unchanged by PCM. Thus, mild PCM significantly increases macrophage-PGE2 production secondary to increased AA phospholipid content, with subsequent inhibition of O2- and MFR expression. Severe PCM inhibits macrophage (O2-) through depletion of critical membrane phospholipid components with subsequent impairment in signal transduction.     

Impaired macrophage function in severe protein-energy malnutrition.

Protein-energy malnutrition induces immunosuppression that predisposes to sepsis, but the mechanisms are unclear. This study examines the role of the macrophage in host defense in the malnourished state. Swiss-Webster mice (N = 300) were randomly allocated to control (24% casein) or low-protein (2.5% casein) diets for 8 weeks. We studied the ability of two populations of macrophages, peritoneal macrophages, and Kupffer cells to produce superoxide anion after in vivo administration of endotoxin or mycobacterium (bacille Calmette-Guérin). Phorbol diester and Candida albicans were used as stimuli. In another group of mice, we evaluated the ability of interferon gamma to up-regulate superoxide anion release and Candida phagocytosis and killing. Mice under protein-energy malnutrition demonstrated decreased mean body weights, serum protein levels, and cell yields. Superoxide anion production in resident and activated (lipopolysaccharide, interferon gamma, bacille Calmette-Guérin infection) peritoneal macrophages was significantly reduced in the malnourished group. Candida phagocytosis and killing were also both depressed in malnourished mice. Kupffer cells failed to generate superoxide anion in all groups. We conclude that severe protein-energy malnutrition significantly impairs macrophage function, which could diminish response to acute and chronic septic challenges. Interferon gamma up regulated peritoneal macrophage and Kupffer cell microbicidal function, which suggests a therapeutic role for this lymphokine in the malnourished septic host.     

Macrophage effector mechanisms in melanoma in an experimental study.

BACKGROUND: The tumor-bearing state is known to induce immune dysfunction that contributes to increased infectious complications and tumor progression. However, the mechanisms underlying this immunosuppression remain unclear. HYPOTHESIS: Macrophage (MO) dysfunction may play a role in tumor-induced immunosuppression. DESIGN AND MAIN OUTCOME MEASURES: Using a murine model, this study investigated the effects of melanoma growth on peritoneal macrophage effector molecule and prostaglandin production, MO-mediated cytotoxicity, and candidacidal mechanisms. Female C57BL/6 mice were inoculated with 106 B16 melanoma cells or a salt solution subcutaneously. Mice were euthanized 3 weeks later and peritoneal MOs were harvested and assayed for nitric oxide, superoxide anion, tumor necrosis factor alpha, and prostaglandin E(2)production. Macrophage-mediated cytotoxicity against B16 melanoma targets and MO candidacidal mechanisms were also measured. RESULTS: Macrophage production of nitric oxide, superoxide anion, and tumor necrosis factor alpha were significantly decreased, while prostaglandin E(2)production was increased in MOs from melanoma-bearing mice. Concomitantly, MO-mediated cytotoxicity and candidacidal mechanisms were significantly impaired. CONCLUSIONS: Melanoma growth leads to decreased MO effector molecule production, increased prostaglandin E(2)production, and impaired MO cytotoxic and candidacidal mechanisms. These results may help explain the observed increased infectious complications in the tumor-bearing host. Strategies aimed at restoring MO function may have therapeutic potential.     

Intracellular survival of Candida albicans in peritoneal macrophages from chronic peritoneal dialysis patients

Candidal peritonitis is a tenacious infection in patients undergoing chronic peritoneal dialysis. Since little is known about host defenses of the human peritoneal cavity against fungi, we investigated the interaction of peritoneal macrophages (PM phi) from uninfected dialysis patients with Candida albicans blastospores. Chemiluminescence (CL) techniques were used to assess the respiratory burst activity of these cells, and candidacidal activity was evaluated with a fluorochrome microassay. In sharp contrast to peripheral blood polymorphonuclear leukocytes (PMNs) from healthy donors, which gave a brisk luminol-enhanced CL response to opsonized blastospores and killed 35% of cell-associated organisms, PM phi produced barely detectable luminol-enhanced CL and killed only 13% of intracellular Candida. These findings appeared to be associated with a decreased level of myeloperoxidase in PM phi. The mechanism of intracellular survival of C albicans also appeared to be related to relatively poor triggering of superoxide production during phagocytosis of viable blastospores. The CL response of PMNs to C albicans was opsonin-dependent, and peritoneal dialysis effluent was devoid of opsonic activity. These studies suggest that local cellular and humoral mechanisms of defense are inadequate for protection of peritoneal dialysis patients against candidal peritonitis.     

A murine model of invasive aspergillosis: variable benefit of interferon-gamma administration under in vitro and in vivo conditions

BACKGROUND: Interferon-gamma modulates host defense in a number of infectious diseases. Previous studies have shown that systemic administration of interferon-gamma (IFN-gamma) can enhance survival in experimental invasive aspergillosis (IA). METHODS: Using a novel model of murine IA that is characterized by primary pulmonary infection, we investigated the role of IFN-gamma in the phagocytosis and killing of Aspergillus fumigatus by murine neutrophils and pulmonary alveolar macrophages in vitro and the impact of systemic and regional administration of IFN-gamma on the course of IA in glucocorticoid-treated mice. RESULTS: In vitro, IFN-gamma significantly enhanced phagocytosis and killing function of both neutrophils and alveolar macrophages from normal animals, but not cortisone-treated animals. In vivo, intravenous administration of IFN-gamma did not improve phagocyte recruitment, in vivo killing, or mortality from IA. Regional (intranasal) administration of IFN-gamma to the lungs enhanced recruitment of phagocytic cells to the lungs and improved in vivo killing, but did not alter (and actually worsened) mortality from IA. CONCLUSIONS: The in vitro and in vivo effects of IFN-gamma in IA are contingent on many variables, including the route of administration and the specific pathogenesis of infection.     

The urokinase plasminogen activator receptor is crucially involved in host defense during acute pyelonephritis

The urokinase plasminogen activator receptor (uPAR) is expressed at the cell surface of inflammatory cells and plays an important role in neutrophil migration. To investigate the in vivo role of uPAR during urinary tract infection, acute pyelonephritis was induced in uPAR-/- and wild-type (WT) mice by intravesical inoculation with 1 x 10(9) colony-forming units (CFU) of uropathogenic Escherichia coli. Mice were killed after 24 and 48 h, after which bacterial outgrowth and cytokine levels in kidney homogenates were determined. Influx of neutrophils was quantified by myeloperoxidase-enzyme-linked immunosorbent assay. uPAR-/- kidneys had significantly higher numbers of E. coli CFU, accompanied by higher levels of interleukin-1beta (IL-1beta), IL-6, keratinocyte-derived chemokine (KC), macrophage inflammatory protein-2 (MIP-2), and tumor necrosis factor-alpha (TNF-alpha). However, the number of infiltrating neutrophils was similar in uPAR-/- and WT mice at both time points, suggesting that uPAR-/- neutrophils have a lower ability to eliminate E. coli. To further investigate this, neutrophil oxidative burst and phagocytosis was measured. The generation of reactive oxygen species upon stimulation with E. coli was not diminished in uPAR-/- neutrophils compared with WT. Interestingly, uPAR-/- neutrophils displayed significantly impaired phagocytosis of E. coli organisms compared with WT neutrophils. We conclude that uPAR is crucially involved in host defense through phagocytosis during E. coli induced acute pyelonephritis.     

Endotoxin promotes synergistic lethality during concurrent Escherichia coli and Candida albicans infection.

Previous studies have suggested that the lipopolysaccharide (LPS, endotoxin) component of the gram-negative bacterial cell wall is a key virulence factor that serves to enhance mortality during infections in which fungi and gram-negative bacteria are copathogens. To test this hypothesis, mice were challenged ip with Escherichia coli 0111:B4, Candida albicans, or both, and the effect of administration of anti-E. coli 0111:B4 LPS monoclonal antibody (mAb) 8G9 on endotoxemia, bacteremia, and mortality was assessed. E. coli (2 x 10(7) colony-forming units (CFU)) plus C. albicans (6 x 10(7) CFU) infection produced 100% mortality at 7 days, compared to the relatively low mortality caused by infection with either E. coli or C. albicans alone (20 and 3%, respectively, P less than 0.01). Administration of mAb 8G9 to animals receiving both pathogens reduced mortality (100% versus 14%, P less than 0.05), endotoxemia (3653 +/- 3187 versus 2 +/- 2 endotoxin units (EU), P less than 0.01), and bacteremia (4.2 +/- 2.3 versus 1.1 +/- 2.1 log(CFU/ml), P less than 0.01) compared to animals receiving saline alone. In a separate series of experiments, purified E. coli 0111:B4 LPS was administered in place of viable E. coli. The simultaneous injection of 200 micrograms E. coli LPS and C. albicans (6 x 10(7) CFU) produced 93% mortality at 7 days, compared to the low mortality that occurred following injection with either E. coli 0111:B4 LPS or C. albicans alone (21 and 3% respectively, P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Bactericidal activity against coagulase-negative staphylococci is impaired in infants receiving long-term parenteral nutrition

OBJECTIVE: To examine the role of total parenteral nutrition (TPN) in predisposing infants to infection caused by coagulase-negative staphylococci. SUMMARY BACKGROUND DATA: Total parenteral nutrition is an important means of providing essential nutrients to newborn infants. However, its use has been associated with complications, particularly infection caused by coagulase-negative staphylococci. Recent data suggest that TPN may modulate immune function; however, reports directly indicating impaired immunity against coagulase-negative staphylococci during TPN are limited. METHODS: Study 1 involved 31 infants younger than 4 months who had undergone surgery and were not receiving antibiotics; 20 were receiving TPN and 11 were receiving a normal enteral diet. An in vitro whole blood model was used to measure the host bactericidal activity against coagulase-negative staphylococci. Bacterial killing and phagocytosis were measured after a 45-minute challenge with viable coagulase-negative staphylococci. In study 2, whole blood killing and intracellular killing of coagulase-negative staphylococci were measured in five newborn infants (younger than 2 months) who were receiving long-term TPN (>10 days), five control infants receiving a normal enteral diet, and five healthy adults. RESULTS: In study 1, infants receiving a normal enteral diet showed a high capacity to ingest and kill coagulase-negative staphylococci. In contrast, the blood of infants receiving long-term TPN showed a reduction in coagulase-negative staphylococci phagocytosis and killing. There were significant negative linear correlations between the duration of TPN and killing of coagulase-negative staphylococci and phagocytosis of coagulase-negative staphylococci. In study 2, infants receiving long-term TPN had lower whole blood killing and intracellular killing than infants receiving a normal enteral diet and healthy adult volunteers. These data seem to indicate a neutrophil dysfunction mediated by TPN in infancy. CONCLUSIONS: Host defense mechanisms, including phagocytosis and killing of coagulase-negative staphylococci, are impaired during long-term TPN. The impaired bactericidal activity seems to be related to defective intracellular killing in neutrophils. These findings may explain the high rate of septicemia caused by coagulase-negative staphylococci in infants receiving TPN.     

Malnutrition-induced macrophage apoptosis

BACKGROUND: Human and murine studies suggest protein-calorie malnutrition (PCM) results in significant host immunosuppression resulting in increased morbidity and mortality. Apoptosis has been implicated as an important mediator in the immunosuppression observed in several disease states. This study was designed to characterize macrophage apoptosis in a murine model of PCM and investigate components that regulate the apoptotic process, such as protein kinase C (PKC) and Bcl-2 activity. METHODS: Swiss-Webster mice (n = 50) were randomly assigned to receive either a control (24% protein) or a PCM diet (0% protein) for 7 days. Peritoneal macrophages were harvested and detection of apoptosis was performed by terminal deoxy-transferase-mediated deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL) and propidium iodide DNA staining under baseline and pro-apoptotic conditions. Pro-apoptotic conditions included cells treated with tumor necrosis factor-alpha (TNF-alpha) (10 ng/mL), interferon-gamma (IFN-gamma) (10 ng/mL), and a combination of both agents. In addition, levels of PKC activity and expression of Bcl-2 and p53 protein were measured. RESULTS: Peritoneal macrophages from PCM mice had a significantly greater amount of apoptosis at baseline and under stimulated conditions compared with controls. Levels of PCM apoptosis were elevated at baseline by TUNEL staining compared with macrophages from the control group (16.5% +/- 1.4%, versus 4.5% +/- 1.1%, P <.01). In addition, peritoneal macrophages from the malnourished animals were significantly more susceptible to the apoptotic effect of TNF-alpha and the effects of INF-gamma (27.3% +/- 2.1% and 31% +/- 1.4%) compared with control mice (5.5% +/- 0.7% and 7.2% +/- 0.5%, P <.01), respectively. Again, an increase in the baseline apoptosis rate was demonstrated in peritoneal macrophages from PCM mice compared with control fed mice (13.2% +/- 4.4% versus 4.3% +/- 3.1%, P <.01) as measured by propidium iodide staining. The combination of agents, TNF-alpha and INF-gamma, resulted in an additive apoptotic effect in the malnourished host compared with the control animals (43.4% +/- 4.7% versus 10.5% +/- 2.2%, P <.01), respectively. Furthermore, there was a significant decrease in the mean total PKC activity in the malnourished macrophages compared with results in controls (110,000 +/- 8000 versus 60,000 +/- 4000 cpm, P <.01). Similar changes were also observed in PKC cytosolic and membrane activity between both groups. In addition, Bcl-2 protein expression was significantly decreased in PCM animals compared with control animals. CONCLUSIONS: Thus, peritoneal macrophages from PCM mice exhibit significantly greater levels of apoptosis at baseline and when stimulated with pro-apoptotic agents compared with controls. The propensity of macrophages from PCM mice to undergo apoptosis may be attributable in part to decreased PKC activity and Bcl-2 protein expression. These findings may help to explain the associated immune dysfunction observed in malnutrition.     

Anti-neoplastic effects of interleukin-4.

Interleukin-4 (IL-4) is a cytokine, with potential anti-neoplastic effects. This study examined the effects of IL-4 on host anti-tumor responses in a murine model. C57/B16 mice (n = 40) were randomized to receive Lewis lung carcinoma (10(6) cells: right flank; sc) or saline, and sacrificed 10 days postinoculation for assessment of peritoneal macrophage (PMO) anti-tumor mechanisms [superoxide anion generation (O2-), tumor necrosis factor (TNF), and TNF-independent (P815) cytotoxicity], splenocyte mixed lymphocyte response (MLR) (Balb/c stimulator), and cytotoxic lymphocyte generation (CTL against P815). Cells were cultured +/- IL-4 (100 U/ml). In a second study, 20 mice received Lewis lung implants (sc) and were randomized on Day 21 to receive daily IL-4 (1000 U/mouse; ip) or saline. Tumor volumes and median survival were assessed. Tumor necrosis factor-independent cytotoxicity (O2-, MLR and CTL) was impaired in the tumor-bearing (TB) study group. Interleukin-4 administered to cultured cells from TB mice enhanced O2-, as well as MLR and CTL (P less than 0.01), and decreased TNF release but did not alter PM phi TNF-independent anti-tumor responses (P815). In vivo administration of IL-4 significantly decreased tumor growth (P less than 0.05) after 10 days of treatment and significantly prolonged median host survival (P less than 0.05). These findings indicate the therapeutic potential of IL-4 in the TB host which may function through downregulation of TNF production while potentiating certain T cell-dependent and independent anti-tumor immune mechanisms.     

Granulocyte macrophage colony-stimulating factor improves survival in two models of gut-derived sepsis by improving gut barrier function and modulating bacterial clearance.

OBJECTIVE: The effect of recombinant murine granulocyte macrophage colony-stimulating factor (rmGM-CSF) on survival and host defense was studied using two clinically relevant models of infection that included transfusion-induced immunosuppression. SUMMARY BACKGROUND DATA: Granulocyte macrophage colony-stimulating factor improves resistance in several models of infection, but its role in transfusion-induced immunosuppression and bacterial translocation (gut-derived sepsis) has not been defined. METHODS: Balb/c mice were treated with 100 ng of rmGM-CSF or placebo for 6 days in a model of transfusion, burn, and gavage, or cecal ligation and puncture (CLP). Translocation was studied in the first model. RESULTS: Survival after transfusion, burn, and gavage was 90% in rmGM-CSF-treated animals versus 35% in the control group (p < 0.001). After CLP, survival was 75% in the rmGM-CSF group versus 30% in the control group (p = 0.01). Less translocation and better killing of bacteria was observed in the tissues in animals treated with rmGM-CSF. CONCLUSION: The ability of rmGM-CSF to improve gut barrier function and enhance killing of translocated organisms after burn injury-induced gut origin sepsis was associated with improved outcome. Granulocyte macrophage colony-stimulating factor also improved survival after CLP.     

Interleukin 1 and its relationship to endotoxin tolerance.

Endotoxin (lipopolysaccharide [LPS])-induced cytokine release has been implicated in the pathogenesis of sepsis. Sublethal doses of LPS induce tolerance to a septic insult. This study evaluated pretreatment with interleukin 1 (IL-1) against an LPS challenge and examined its relationship to endotoxin tolerance. C3H/HeN mice (N = 100) were injected intraperitoneally with phosphate-buffered saline (control group), IL-1 (200 micrograms/kg), or LPS (1 mg/kg) for 3 days. On day 5, peritoneal macrophages were harvested and assayed for antimicrobial activity (superoxide anion production and Candida albicans phagocytosis). Serum cytokine levels and survival after an LPS challenge on day 5 were also assessed. Pretreatment with IL-1 or LPS significantly increased superoxide anion production, C albicans phagocytosis, and survival compared with pretreatment with phosphate-buffered solution. Interleukin 6 levels significantly decreased in the IL-1 and LPS groups. Peak levels of tumor necrosis factor significantly decreased only in the LPS group. Thus, pretreatment with IL-1 or low doses of LPS may exert protective effects by decreasing levels of interleukin 6 while increasing antimicrobial activity. Mice pretreated with IL-1 were protected from endotoxin despite elevated peak levels of tumor necrosis factor, suggesting a different mechanism for endotoxin tolerance than for tolerance to tumor necrosis factor.     

Fibrin in peritonitis. V. Fibrin inhibits phagocytic killing of Escherichia coli by human polymorphonuclear leukocytes.

Fibrin deposition initiated by peritonitis is thought to be an important local defense mechanism because it sequesters and walls off bacterial spillage. However, fibrin has been shown to predispose to residual abscess formation in rat peritonitis model. To examine the potential mechanisms of this effect, fibrin was tested in vitro for its inhibitory effect on neutrophil function. At all concentrations tested (50-1000 mg/dl), fibrin significantly impaired the ability of neutrophils to kill Escherichia coli. This inhibition occurred in a dose dependent fashion with almost complete prevention of killing at the highest concentration tested. Further studies showed that pre-exposure to fibrin did not reduce the neutrophil's ability to degranulate, undergo a respiratory burst, or kill E. coli, indicating that fibrin did not cause irreversible damage to the normal microbicidal functions of the neutrophil. However, fibrin, at physiologic concentrations, significantly impaired phagocytosis of radiolabeled E. coli. The data support the concept that phagocytosis of bacteria is impaired by neutrophils enmeshed in fibrin. Thus, contaminated fibrin could act as a nidus for residual abscesses formation following peritonitis even if an adequate number of normal leukocytes were present.     

Effects of ascorbate on leucocytes: Part II. Effects of ascorbic acid and calcium and sodium ascorbate on neutrophil phagocytosis and post-phagocytic metabolic activity.

The effects of ascorbic acid and calcium and sodium ascorbate at a concentration range of 10(-6)M - 10(-1)M on polymorphonuclear leucocyte (PMN) phagocytosis of Candida albicans and post-phagocytic nitroblue tetrazolium (NBT) reduction, hexose monophosphate shunt (HMS) activity and myeloperoxidase-mediated iodination of ingested protein were investigated. Phagocytosis of C. albicans was unaffected by ascorbate concentrations of 10(-6)M - 10(-2)M; however, progressive inhibition was observed at concentrations of 10(-2)M upwards. Enhancement of resting and stimulated HMS activity and NBT reduction was evident at ascorbate concentrations of 10(-5) M - 10(-2)M. The stimulations of HMS activity and NBT reduction was independent of myeloperoxidase iodination of ingested protein and this latter function was strongly inhibited by ascorbate. Concentrations of ascorbic acid and calcium and sodium ascorbate which caused inhibition of phagocytosis and HMS activity were the same as those which mediated stimulation of cell motility, indicating that independent cellular mechanisms may govern motility and phagocytosis.     

Patterns of bacterial translocation in experimental biliary obstruction

INTRODUCTION: Biliary obstruction is associated with impaired intestinal barrier function and translocation of enteric bacteria to the systemic circulation. Traditional live culture techniques may overlook translocation of dead bacterial fragments that stimulate the inflammatory response. The aim of this study was to estimate the extent and pattern of bacterial translocation in experimental biliary obstruction. MATERIALS AND METHODS: Thirty 9-week-old male Wistar rats were randomized to undergo bile duct ligation (BDL, n = 20) or sham operation (n = 10). Seven days after operation, each animal received 1 ml of (111)indium-oxyquinolone-labeled Escherichia coli p.o. Samples of liver, spleen, mesenteric lymph nodes, and lung were harvested 4 h later and analyzed for live bacteria and (111)indium activity. RESULTS: There was significantly more live bacterial translocation detected in BDL animals than in sham-operated animals (P = 0.00008, chi(2)). Labeled bacterial fragments were detected in all locations sampled in all animals. Sham-operated animals had significantly more labeled bacterial fragments detected in the liver (P = 0.0001) and the spleen (P = 0.03) than the BDL animals. The mean total bacterial survival in the BDL group was 30 +/- 13% and 0% in the sham operated group. CONCLUSION: These results demonstrate that non-viable bacterial fragments are present in sterile extra-intestinal sites in normal animals and that translocation of live bacteria is markedly increased in experimental biliary obstruction. These results also suggest that failure of bacterial killing is an important factor facilitating bacterial translocation in the presence of established biliary obstruction.     

Impaired nonspecific cellular immunity in experimental cholestasis.

The abilities of polymorphonuclear leukocytes (PMN) and pulmonary alveolar macrophages (PAM), to demonstrate chemotaxis, phagocytosis, and superoxide release after bile duct ligation in the rat were investigated to determine the effect of cholestasis on nonspecific cellular immune mechanisms. Chemotactic response to C5a and FMLP, phagocytosis of 14C labeled Staphylococcus aureus, and zymosan-induced superoxide release were evaluated 21 days after bile duct ligation (BDL), sham operation, or in normal controls. Serum total bilirubin level was elevated after BDL (p less than 0.01). Chemotactic ability was similar to each group. PMN phagocytic uptake of 14C labeled Staphylococcus aureus was depressed in BDL (p less than 0.05). BDL rats exhibited impaired PAM phagocytic indices and improved PMN superoxide release (p less than 0.03). PAM superoxide release was similar in each study group. Alterations in phagocytic function with cholestasis are important deficits in nonspecific cellular immunity that may contribute to the high incidence of infective complications associated with obstructive jaundice.     

CD4+ T-cell-dependent immune damage of liver parenchymal cells is mediated by alloantibody

BACKGROUND: Allogeneic hepatocytes initiate both CD4- and CD8-dependent rejection responses. The current studies address the hypothesis that acute damage of allogeneic liver parenchymal cells by the CD4-dependent pathway is alloantibody-mediated and examines immune conditions which promote activation of this pathway. METHODS: The role of alloantibody in CD4-dependent hepatocyte rejection was evaluated by assessing hepatocyte (FVB/N, H-2q) survival in CD8-depleted B-cell knockout (KO) (H-2b) recipients and by monitoring hepatocyte survival in C57BL/6.SCID (H-2b) recipients transfused with donor-reactive alloantibody. The development of donor-reactive alloantibody in C57BL/6 (H-2b), CD8-depleted C57BL/6, CD8 KO (H-2b), IFN-gamma KO (H-2b), perforin KO (H-2b), and FasL mutant gld/gld (H-2b) hepatocyte recipients was assessed. RESULTS: Hepatocyte rejection in B-cell KO mice was significantly delayed by CD8+ T-cell depletion (median survival time [MST], 35 days) when compared to untreated (MST, 8 days) and CD4-depleted (MST, 10 days) recipient mice. Transfusion of donor-reactive alloantibody into SCID recipients with functional hepatocellular allografts was sufficient to precipitate rejection in a dose-dependent fashion. Donor-reactive alloantibody was minimal in the serum of C57BL/6 hepatocyte recipients, but was produced in significant quantities in hepatocyte recipients genetically deficient in or depleted of CD8+ T cells and in recipients with impaired cytotoxic effector mechanisms. In addition, recipients with defects in Th1 immunity, such as IFN-gamma KO recipients, also produced readily detectable alloantibody. CONCLUSIONS: Collectively, these data support the hypothesis that acute immune damage of allogeneic hepatocytes by the CD4-dependent pathway is mediated by alloantibody and that this pathway is favored when Th1- or cell-mediated cytotoxic effector immune mechanisms are impaired.