Synbiotics decrease the incidence of septic complications in patients with severe SIRS: a preliminary report.

The purpose of this study was to evaluate if synbiotic therapy can correct the deteriorated gut flora and environment in patients with severe systemic inflammatory response syndrome (SIRS). Twenty-nine SIRS patients, who fulfilled a serum C-reactive protein (CRP) level >10 mg/dl, received synbiotics (Bifidobacterium breve, Lactobacillus casei, and galactooligosaccharides) (S group) and were compared with previous observations in 26 patients without synbiotics (NS group). Analysis of fecal flora confirmed that patients in the S group had significantly greater levels of beneficial Bifidobacterium, Lactobacillus, and total organic acids (particularly short-chain fatty acids) than those in the NS group. The incidence of infectious complications such as enteritis, pneumonia, and bacteremia was significantly lower in the S group than in the NS group. Synbiotics maintain the gut flora and environment and decrease the incidence of septic complications in patients with severe SIRS. Further randomized controlled study is necessary to determine the effects of synbiotics.

Early activation of gammadelta T lymphocytes in patients with severe systemic inflammatory response syndrome.

Innate immunity plays an important role in host defense after severe insult. gammadelta T lymphocytes are recognized as the first line of defense against microbial invasion. In this study, we evaluated gammadelta T lymphocytes in the peripheral blood of patients with severe systemic inflammatory response syndrome (SIRS), and examined on role of these cells. Thirty-seven patients with severe SIRS (SIRS criteria and serum C-reactive protein > or = 10 mg/dL) and 27 healthy volunteers were studied. Severe SIRS was caused by trauma in 14 patients (Injury Severity Score of 30.1 +/- 10.8) and by sepsis in 23 patients. The counts of gammadelta and alphabeta T lymphocytes were determined by flow cytometry of cells stained with monoclonal antibodies to gammadelta and alphabeta T lymphocyte receptors. The activation of these cells was evaluated by flow cytometry of cells stained with monoclonal antibodies to CD69 and HLA-DR. Serial counts and activation of gammadelta and alphabeta T lymphocytes were also determined in eight trauma patients (Injury Severity Score of 31.0 +/- 13.5) during a 2-week observation period. The count of gammadelta T lymphocytes in the peripheral blood of SIRS patients (30.1 +/- 6.0/microL) was significantly lower (P < 0.05) than that of the healthy volunteers (104.3 +/- 10.9/microL). The expression of CD69, an index of early activation of T lymphocytes, was significantly greater on gammadelta T lymphocytes from SIRS patients (patients 23.9% +/- 3.4%, healthy controls 4.8% +/- 0.6%, P < 0.05). In trauma patients, the expression of CD69 on gammadelta T lymphocytes increased rapidly within 48 h after injuries. In conclusion, gammadelta T lymphocytes are activated and decreased in the peripheral blood of severe SIRS patients. In trauma patients, the activation of gammadelta T lymphocytes occurs in the fairly acute phase after injuries. These results suggest a significant role for gammadelta T lymphocytes as early responders after severe insult.

A neutrophil elastase inhibitor, sivelestat, improves leukocyte deformability in patients with acute lung injury.

BACKGROUND: The objective of this study was to evaluate whether the neutrophil elastase (NE) inhibitor, sivelestat, improves leukocyte deformability and pulmonary function in patients with acute lung injury (ALI). PATIENTS AND METHODS: Twenty-four patients with systemic inflammatory response syndrome (SIRS) were divided into two groups: those with ALI (ALI group, n = 14), and those without ALI (non-ALI group, n = 10). Within 72 hours after the diagnosis, we measured the total leukocyte count (TLC), C-reactive protein (CRP) level, NE concentration, APACHE II score, Goris multiple organ failure (MOF) index, respiratory index (RI), lung injury score (LIS), and oxygenation index (P/F ratio). Leukocyte deformability was examined with a microchannel array etched on a single-crystal silicon tip that simulates the microvasculature. The number of obstructed microchannels (NOM) because of stiffened neutrophils and transit time (TT), defined as the time needed for 100 microL of whole blood to pass through the microchannels, were determined. We then administered sivelestat (4.8 mg/kg/d) to nine ALI patients (sivelestat group) for 5 days and compared with seven ALI patients treated previously without sivelestat (conventional group). The factors described above were measured before and 5 days after treatment. RESULTS: There were no significant differences in age, TLC, CRP, APACHE II score, and MOF index between ALI and non-ALI group. RI and LIS were higher and the P/F ratio was significantly lower in the ALI group than in the non-ALI group. NE concentration, NOM, and TT were significantly higher in the ALI group than in the non-ALI group (p < 0.05). After 5 days of treatment with sivelestat, the APACHE II score, MOF index, RI, LIS, NE concentration, TT, and NOM were lower and the P/F ratio was significantly higher than baseline values and those in the conventional group (p < 0.05). CONCLUSION: NE concentration and neutrophil rigidity are significantly increased in SIRS patients with ALI. Sivelestat appears to reduce NE concentration and neutrophil stiffness and improve pulmonary oxygenation in patients with ALI.

Enhanced expression of intracellular heme oxygenase-1 in deactivated monocytes from patients with severe systemic inflammatory response syndrome.

BACKGROUND: Monocyte deactivation is an important contributor to infectious susceptibility in critically ill patients. However, the mechanism of monocyte deactivation has not been fully elucidated. Recently, intracellular heme oxygenese-1 (HO-1), an anti-inflammatory heat-shock protein, was reported to be activated by Toll-like receptors (TLRs), and to inhibit inflammatory cytokine production such as that of TNF-alpha. In the present study, we evaluated the expression of intracellular HO-1 and TLRs in monocytes from patients with severe systemic inflammatory response syndrome (SIRS) and examined the role of HO-1 in monocyte deactivation. PATIENTS: Twenty-seven patients who fulfilled the criteria for severe SIRS and had a serum C-reactive protein (CRP) level >10 mg/dL were included in this study. The cause of SIRS was sepsis in 16 patients, trauma in 7, and other in 4. Expression of intracellular HO-1, surface TLR2 and TLR4, and intracellular cytokines (TNF-alpha, Interleukin-6) stimulated via TLR activation were measured in circulating monocytes by flow cytometry. Intracellular HO-1 expression was evaluated in normal monocytes stimulated with patient serum. Serum cytokine levels were also measured. Patient data were compared with data from healthy volunteers (n = 16). RESULTS: Cytoplasmic HO-1 was clearly detected by fluorescence microscopy. Expression of HO-1, TLR2, and TLR4 in monocytes was significantly enhanced in patients with severe SIRS compared with that in healthy volunteers, whereas intracellular TNF-alpha expression with peptidoglycan was significantly decreased (p < 0.05) in patients compared with that in healthy volunteers. HO-1 expression was significantly enhanced in normal monocytes stimulated with patient serum. Intracellular HO-1 levels were positively related to serum TNF-alpha levels in patients (r = 0.46). CONCLUSIONS: Expression of intracellular HO-1 and of TLRs was enhanced in deactivated monocytes from patients with SIRS. Increased production of intracellular HO-1 in response to serum factors may play a role in monocyte deactivation after systemic inflammation.

Serial changes in leukocyte deformability and whole blood rheology in patients with sepsis or trauma.

BACKGROUND: The objective of this study was to investigate serial changes in leukocyte deformability and rheologic properties of whole blood in patients with sepsis or trauma. METHODS: Seventeen sepsis patients and 22 trauma patients were enrolled. Leukocyte deformability and rheologic properties of whole blood were determined with the use of a microchannel array etched on a single-crystal silicon tip, simulating the microvasculature. The number of obstructed microchannels (NOM) was used as a measure of leukocyte deformability. Transit time (TT), i.e., the time taken for 100 microL of whole blood to pass through the microchannel array was also used as rheologic measure. Oxidative activity and F-actin content of neutrophils was measured in patients with sepsis. RESULTS: NOM and TT significantly increased in patients when sepsis was diagnosed. In survivors, NOM and TT decreased at the time of recovery from sepsis, but in non-survivors values remained high. Oxidative activity and F-actin content of neutrophils increased significantly as leukocyte deformability decreased. In patients with severe trauma, NOM and TT increased after injury and decreased by the time of recovery from the critical stage. CONCLUSION: We conclude that leukocyte deformability decreases in patients with sepsis or severe trauma and that this change negatively affects rheologic properties of whole blood.

The balance between expression of intranuclear NF-kappaB and glucocorticoid receptor in polymorphonuclear leukocytes in SIRS patients.

BACKGROUND: We previously reported enhanced expression of nuclear factor kappa B (NF-kappaB) in activated polymorphonuclear leukocytes (PMNLs) from patients with systemic inflammatory response syndrome (SIRS). Inflammatory response, however, is not regulated only by stimulatory transcription factors. Glucocorticoid receptor (GR) has been recently reported to play an important role in anti-inflammatory signal transduction. The objective of our study was to evaluate the balance between expression of intranuclear NF-kappaB and GR in PMNLs from SIRS patients. METHODS: In study 1, 29 patients with severe SIRS, who fulfilled the criteria for SIRS and had a serum C-reactive protein level of more than 10 mg/dL, were included. Expression of intranuclear NF-kappaB and GR in PMNLs was measured by flow cytometry with antibodies specific for NF-kappaB subunit p65 and GR. PMNL oxidative activity and serum cytokine levels were also measured. Study 2 included 13 patients with severe trauma (Injury Severity Score 24.6 +/- 12.2). We measured serial changes in expression of intranuclear NF-kappaB and GR in days 0 to 2, 3 to 6, and 7 to 14 after injury. RESULTS: In study 1, expression of both intranuclear NF-kappaB and GR in PMNLs was significantly higher in SIRS patients than in healthy controls. There was a strong correlation between expression of these two transcription factors (r = 0.78). Positive correlations were also found between PMNL oxidative activity and both transcription factors. In study 2, expression of both NF-kappaB and GR in PMNLs was markedly elevated on days 3 to 6 after injury and changed serially with strong mutual correlation. CONCLUSIONS: In activated PMNLs from SIRS patients, levels of both intranuclear NF-kappaB and GR were elevated and strongly correlated. In trauma patients, NF-kappaB and GR in PMNLs changed serially with strong mutual correlation. Further studies are needed to clarify the effect of the balance of NF-kappaB and GR on PMNL activation and systemic inflammatory process.

Hepatocyte growth factor in polymorphonuclear leukocytes is increased in patients with systemic inflammatory response syndrome.

BACKGROUND: Hepatocyte growth factor (HGF) has a significant effect on the regeneration of epithelial and endothelial cells. Studies have also shown an important role of HGF in wound healing and organ regeneration. Because recent studies indicate that polymorphonuclear leukocytes (PMNLs) store HGF in their specific granules and that HGF can be degranulated in the inflammatory tissue in which activated PMNLs migrate, we evaluated the storage and release of HGF in PMNLs from patients with systemic inflammatory response syndrome (SIRS) and attempted to examine the role of HGF from PMNLs in the systemic inflammatory process. METHODS: Twenty-four patients with SIRS (serum C-reactive protein, 20.2 +/- 12.4 mg/dL [mean +/- SD]) and 18 healthy volunteers were studied. HGF in PMNLs was measured by flow cytometry by using a monoclonal antibody to HGF. The oxidative activity in PMNLs was also measured by flow cytometry. Serum HGF, interleukin (IL)-6, and IL-8 levels in each patient were measured by enzyme-linked immunosorbent assay. HGF degranulation from PMNLs was evaluated in 10 patients. RESULTS: Immunocytochemistry under fluorescence microscopy revealed enhanced expression of HGF in the granules of PMNLs. HGF in PMNLs significantly increased in patients with SIRS compared with PMNLs from healthy volunteers (SIRS, 171.0 +/- 6.6 fluorescence/cell; control, 130.7 +/- 3.8 fluorescence/cell). N-formylmethionyl-leucyl-phenylalanine and lipopolysaccharide stimulation induced further increase of HGF fluorescence in PMNLs from patients. HGF degranulation from PMNLs was also significantly enhanced in patients. Moreover, oxidative activity in PMNLs was significantly enhanced in patients with SIRS. Plasma HGF (pHGF) correlated positively with IL-6 and IL-8 levels in patients (pHGF and IL-6, gamma = 0.635, p < 0.05; pHGF and IL-8, gamma = 0.827, p < 0.01), but these values did not correlate with HGF in PMNLs. CONCLUSION: Activated PMNLs in SIRS patients increased HGF in their granules and demonstrate enhanced degranulation of HGF. The release of HGF from migrated PMNLs in the inflammatory tissue may play an important role in wound healing and organ regeneration under those conditions.

Enhanced production of endothelial microparticles with increased binding to leukocytes in patients with severe systemic inflammatory response syndrome.

BACKGROUND: The vascular endothelium sustains substantial damage after severe insult. Recently, activated endothelial cells have been reported to produce microparticles in vitro. The objective of this study was to evaluate endothelial microparticle formation and microparticle-leukocyte interaction among patients with severe systemic inflammatory response syndrome (SIRS). METHODS: The participants in this study were 28 patients with severe SIRS (SIRS criteria and serum C-reactive protein > 10 mg/dL) and 18 healthy volunteers. Endothelial microparticles in the blood, microparticle-polymorphonuclear leukocyte (PMNL) binding, and PMNL oxidative activity were measured by flow cytometry. Soluble E-selectin, thrombomodulin, and plasminogen activator inhibitor-1 levels in the blood, variables representing systemic vascular endothelial cell activation and damage, and coagulative activity in the blood also were measured. RESULTS: Endothelial microparticle levels in the blood, microparticle binding to PMNLs, and oxidative activity in PMNLs increased significantly in patients with severe SIRS, as compared with the values in healthy volunteers. Soluble E-selectin, thrombomodulin, plasminogen activator inhibitor-1, and procoagulant activity in the blood also increased in these patients. CONCLUSIONS: Activated vascular endothelial cells with increased procoagulant activity enhance production of microparticles with increased binding to leukocytes in patients with severe SIRS. Endothelial microparticles may be involved in the pathogenesis of endothelial injury after severe insult.

Paradoxical cytoskeleton and microparticle formation changes in monocytes and polymorphonuclear leukocytes in severe systemic inflammatory response syndrome patients.

BACKGROUND: Circulating monocytes and polymorphonuclear leukocytes (PMNLs) are considered as central regulators controlling systemic inflammatory response after severe insults. Recently, activated monocytes and PMNLs have been reported to produce microparticles (MPs) in vitro. The objective of this study was to evaluate production of MPs and changes of cytoskeleton in monocytes from severe systemic inflammatory response syndrome (SIRS) patients, and to compare them with those in PMNLs. METHODS: Twenty severe SIRS patients (SIRS criteria and serum C-reactive protein > 10 mg/dL) and 15 healthy volunteers were included. MP formation and F-actin content in monocytes and PMNLs were measured by flow cytometry in the presence or absence of lipopolysaccharide or formylmethionyl-leucyl-phenylalanine (FMLP). The membrane expression of human leukocyte antigen-DR and CD64 in monocytes and O2- production in PMNLs were also measured by flow cytometry. RESULTS: In severe SIRS patients, MP formation with and without lipopolysaccharide in monocytes significantly decreased in comparison with those in normal controls (p < 0.05), whereas those with and without FMLP in PMNLs increased (p < 0.05). F-actin content with and without FMLP in monocytes also significantly decreased in patients (p < 0.05), whereas those in PMNLs increased as compared with normal controls (p < 0.05). The expression of human leukocyte antigen-DR in monocytes significantly decreased in patients (p < 0.05), which indicated monocyte modulation. The O2- production in PMNLs increased in patients (p < 0.05), which showed PMNL activation. CONCLUSION: The changes of MP formation and cytoskeleton in circulating monocytes and PMNLs were paradoxically different in severe SIRS patients.