Evidence for metabolic endotoxemia in obese and diabetic Gambian women.

OBJECTIVE: Emerging evidence from animal models suggests that translocation of bacterial debris across a leaky gut may trigger low-grade inflammation, which in turn drives insulin resistance. The current study set out to investigate this phenomenon, termed 'metabolic endotoxemia', in Gambian women. METHODS: In a cross-sectional study, we recruited 93 age-matched middle-aged urban Gambian women into three groups: lean (body mass index (BMI): 18.5-22.9 kg m(-2)), obese non-diabetic (BMI: 30.0 kg m(-2)) and obese diabetic (BMI: 30.0 kg m(-2) and attending a diabetic clinic). We measured serum bacterial lipopolysaccharide (LPS) and endotoxin-core IgM and IgG antibodies (EndoCAb) as measures of endotoxin exposure and interleukin-6 (IL-6) as a marker of inflammation. RESULTS: Inflammation (IL-6) was independently and positively associated with both obesity and diabetes (F=12.7, P<0.001). LPS levels were highest in the obese-diabetic group compared with the other two groups (F=4.4, P<0.02). IgM EndoCAb (but not total IgM) was highly significantly reduced in the obese (55% of lean value) and obese diabetic women (30% of lean; F=21.7, P<0.0001 for trend) compared with lean women. CONCLUSION: These data support the hypothesis that gut-derived inflammatory products are associated with obesity and diabetes. Confirmation of these findings and elucidation of the role of the microbiota, gut damage and the pathways for translocation of bacterial debris, could open new avenues for prevention and treatment of type 2 diabetes.

Granulocyte colony-stimulating factor (G-CSF) depresses angiogenesis in vivo and in vitro: implications for sourcing cells for vascular regeneration therapy.

SUMMARY BACKGROUND: The most common source of hematopoietic progenitor cells (HPCs) for hematopoietic reconstitution comprises granulocyte colony-stimulating factor (G-CSF)-mobilized peripheral blood stem cells (PBSCs). It has been proposed that endothelial progenitor cells (EPCs) share precursors with HPCs, and that EPC release may accompany HPC mobilization to the circulation following G-CSF administration. OBJECTIVE: To investigate EPC activity following HPC mobilization, and the direct effects of exogenous G-CSF administration on human umbilical vein endothelial cells (HUVECs) and endothelial outgrowth cells (EOCs), using in vitro and in vivo correlates of angiogenesis. PATIENTS/METHODS: Heparinized venous blood samples were collected from healthy volunteers and from cord blood at parturition. G-CSF-mobilized samples were collected before administration, at apheresis harvest, and at follow-up. PBSCs were phenotyped by flow cytometry, and cultured in standard colony-forming unit (CFU)-EPC and EOC assays. The effect of exogenous G-CSF was investigated by addition of it to HUVECs and EOCs in standard tubule formation and aortic ring assays, and in an in vivo sponge implantation model. RESULTS: Our data show that G-CSF mobilization of PBSCs produces a profound, reversible depression of circulating CFU-EPCs. Furthermore, G-CSF administration did not mobilize CD34+CD133- cells, which include precursors of EOCs. No EOCs were cultured from any mobilized PBSCs studied. Exogenous G-CSF inhibited CFU-EPC generation, HUVEC and EOC tubule formation, microvessel outgrowth, and implanted sponge vascularization in mice. CONCLUSIONS: G-CSF administration depresses both endothelial cell angiogenesis and monocyte proangiogenic activity, and we suggest that any angiogenic benefit observed following implantation of cells mobilized by G-CSF may come only from a paracrine effect from HPCs.

Dissociation of phenotypic and functional endothelial progenitor cells in patients undergoing percutaneous coronary intervention.

OBJECTIVES: Endothelial progenitor cells (EPCs) are circulating mononuclear cells with the capacity to mature into endothelial cells and contribute to vascular repair. We assessed the effect of local vascular injury during percutaneous coronary intervention (PCI) on circulating EPCs in patients with coronary artery disease. DESIGN AND SETTING: Prospective case-control study in a university teaching hospital. PATIENTS: 54 patients undergoing elective coronary angiography. INTERVENTIONS AND MAIN OUTCOME MEASURES: EPCs were quantified by flow cytometry (CD34(+)KDR(+) phenotype) complemented by real-time polymerase chain reaction (PCR), and the colony forming unit (CFU-EC) functional assay, before and during the first 24 hours after diagnostic angiography (n = 27) or PCI (n = 27). RESULTS: Coronary intervention, but not diagnostic angiography, resulted in an increase in blood neutrophil count (p<0.001) and C-reactive protein concentrations (p = 0.001) in the absence of significant myocardial necrosis. Twenty-four hours after PCI, CFU-ECs increased threefold (median [IQR], 4.4 [1.3-13.8] vs 16.0 [2.1-35.0], p = 0.01), although circulating CD34(+)KDR(+) cells (0.019% (SEM 0.004%) vs 0.016% (0.003%) of leucocytes, p = 0.62) and leucocyte CD34 mRNA (relative quantity 2.3 (0.5) vs 2.1 (0.4), p = 0.21) did not. There was no correlation between CFU-ECs and CD34(+)KDR(+) cells. CONCLUSIONS: Local vascular injury following PCI results in a systemic inflammatory response and increases functional CFU-ECs. This increase was not associated with an early mobilisation of CD34(+)KDR(+) cells, suggesting these cells are not the primary source of EPCs involved in the immediate response to vascular injury.

Influence of menstrual cycle on circulating endothelial progenitor cells.

BACKGROUND: Endothelial progenitor cells (EPCs) are circulating mononuclear cells that participate in angiogenesis. The aim of this study was to determine the influence of the menstrual cycle on the number and function of EPCs, and to investigate their relationship with circulating concentrations of sex steroids and inflammatory mediators. METHODS: Ten healthy nulliparous, premenopausal, non-smoking women with regular menses were studied over a single menstrual cycle. Venepuncture was performed in the menstrual, follicular, peri-ovulatory and luteal phases. EPCs were quantified by flow cytometry (CD133(+)CD34(+)KDR(+) phenotype) and the colony-forming unit (CFU-EPC) functional assay. Circulating concentrations of estradiol, progesterone and inflammatory mediators (TNF-alpha, IL-6, sICAM-1 and VEGF) were measured by immunoassays. RESULTS: The numbers of CD133(+)CD34(+)KDR(+) cells were higher in the follicular phase (0.99 +/- 0.3 x 10(6) cells/l) compared with the peri-ovulatory phase (0.29 +/- 0.1 x 10(6) cells/l; P < 0.05). In contrast, the numbers of CFU-EPCs did not vary over the menstrual cycle. There were no correlations between EPCs and concentrations of either circulating sex steroids or inflammatory mediators. CONCLUSIONS: CD133(+)CD34(+)KDR(+) cells but not CFU-EPCs vary during the menstrual cycle. Our findings suggest a potential role for circulating EPCs in the normal cycle of physiological angiogenesis and repair of the uterine endometrium that is independent of circulating sex steroids or inflammatory mediators.

Endotoxin immunity and the development of the systemic inflammatory response syndrome in critically ill children.

BACKGROUND: The systemic inflammatory response syndrome (SIRS) may be triggered by endotoxin. Humans have antibodies directed against the core of endotoxin (endotoxin core antibodies, EndoCAb) that appear to be protective following surgery and in sepsis. We hypothesised that children with elevated antibodies to endotoxin core would be less likely to develop SIRS in their initial period on intensive care. Because of the existing literature we defined two sub-groups according to the primary reason for ICU admission: infection and non-infection. METHODS: We recruited 139 consecutive patients admitted to a paediatric intensive care unit (PICU) with more than one organ failure for longer than 12 h as part of another study. Patients were classified on admission to PICU as having an infectious or a non-infections diagnosis. The occurrence of SIRS within 48 h of admission was recorded along with detailed clinical and demographic data, EndoCAb concentration and the potential confounding variables C-reactive protein and mannose-binding lectin. RESULTS: In the 71 patients admitted without infection (primarily post-operative and head injured) IgG EndoCAb was significantly lower in patients who developed SIRS than those who did not (72 vs. 131 MU/ml), independent of potential confounding variables. In patients with infection there was no significant difference in IgG EndoCAb between children developing SIRS and those who did not (111 vs. 80 MU/ml). CONCLUSION: Head injured and post-operative patients admitted to PICU who develop early SIRS have significantly lower serum IgG EndoCAb levels than those who do not.

Intraoperative oesophageal Doppler guided fluid management shortens postoperative hospital stay after major bowel surgery.

BACKGROUND: Occult hypovolaemia is a key factor in the aetiology of postoperative morbidity and may not be detected by routine heart rate and arterial pressure measurements. Intraoperative gut hypoperfusion during major surgery is associated with increased morbidity and postoperative hospital stay. We assessed whether using intraoperative oesophageal Doppler guided fluid management to minimize hypovolaemia would reduce postoperative hospital stay and the time before return of gut function after colorectal surgery. METHODS: This single centre, blinded, prospective controlled trial randomized 128 consecutive consenting patients undergoing colorectal resection to oesophageal Doppler guided or central venous pressure (CVP)-based (conventional) intraoperative fluid management. The intervention group patients followed a dynamic oesophageal Doppler guided fluid protocol whereas control patients were managed using routine cardiovascular monitoring aiming for a CVP between 12 and 15 mm Hg. RESULTS: The median postoperative stay in the Doppler guided fluid group was 10 vs 11.5 days in the control group P<0.05. The median time to resuming full diet in the Doppler guided fluid group was 6 vs 7 for controls P<0.001. Doppler patients achieved significantly higher cardiac output, stroke volume, and oxygen delivery. Twenty-nine (45.3%) control patients suffered gastrointestinal morbidity compared with nine (14.1%) in the Doppler guided fluid group P<0.001, overall morbidity was also significantly higher in the control group P=0.05. CONCLUSIONS: Intraoperative oesophageal Doppler guided fluid management was associated with a 1.5-day median reduction in postoperative hospital stay. Patients recovered gut function significantly faster and suffered significantly less gastrointestinal and overall morbidity.

A descriptive study of the variation in baseline levels of antiendotoxin core antibodies between US and UK populations.

Low levels of naturally occurring antibodies to the core section of endotoxin (EndoCAb) have been shown to be predictors of poor outcome following major surgery. We performed a retrospective study comparing pre-operative levels in US surgical patients, UK surgical patients and healthy volunteers. Both IgM and IgG EndoCAb levels were higher in the US surgical patients when compared with the other groups (approximately twice as high in the case of IgG EndoCAb). This may reflect genetic or environmental variability between the patient groups, differences in the disease processes, the disparity in the delivery of health care between the two countries or degradation of the samples in transfer.

Calcitonin precursors: early markers of gut barrier dysfunction in patients with acute pancreatitis.

BACKGROUND: Severe acute pancreatitis is associated with an early increase in intestinal permeability and endotoxemia. Endotoxin is a potent stimulator for the production and release of procalcitonin and its components (calcitonin precursors; [CTpr]). The aim of this study is to evaluate the role of plasma CTpr as an early marker for gut barrier dysfunction in patients with acute pancreatitis. METHODS: Intestinal permeability to macromolecules (polyethylene glycol 3350), serum endotoxin and antiendotoxin core antibodies, plasma CTpr, and serum C-reactive protein (CRP) were measured on admission in 60 patients with acute pancreatitis. Attacks were classified as mild (n = 48) or severe (n = 12) according to the Atlanta criteria. RESULTS: Compared with mild attacks of acute pancreatitis, severe attacks were significantly associated with an increase in intestinal permeability index (median: 0.02 vs. 0.006, P < 0.001), the frequency of endotoxemia (73% vs. 41%, P = 0.04), and the extent of depletion of serum IgM antiendotoxin antibodies (median: 43 MMU vs. 100 MMU, P = 0.004). Plasma CTpr levels were significantly elevated in patients with severe attacks compared with mild attacks on both the day of admission and on day 3 (median: 64 vs. 22 fmol/mL, P = 0.03; and 90 vs. 29 fmol/mL, P = 0.003 respectively). A positive and significant correlation was observed between the admission serum endotoxin and plasma CTpr levels on admission (r = 0.7, P < 0.0001) and on day 3 (r = 0.96, P < 0.0001), and between plasma CTpr on day 7 and the intestinal permeability index (r = 0.85, P = 0.0001). In contrast, only a weak positive correlation was observed between peak serum levels of CRP and plasma CTpr on admission (r = 0.3, P = 0.017) and on day 7 (r = 0.471, P = 0.049), as well as between CRP and each of the admission serum endotoxin (r = 0.3, P = 0.03) and the intestinal permeability index (r = 0.375, P = 0.007). CONCLUSIONS: In patients with acute pancreatitis, plasma concentrations of CTpr appear to reflect more closely the derangement in gut barrier function rather than the extent of systemic inflammation.

Hypocalcemia in patients with acute pancreatitis: a putative role for systemic endotoxin exposure.

INTRODUCTION: Hypocalcemia is not uncommon during acute pancreatitis and is associated with a poor outcome. Whilst the mechanisms responsible for its development remain unclear, there is evidence to implicate endotoxemia in other models of sepsis. AIM: To investigate the potential role of systemic endotoxin exposure in the development of hypocalcemia in patients with acute pancreatitis. METHODOLOGY: Adjusted serum calcium was measured daily, and the lowest value within 72 hours of admission for acute pancreatitis was determined. Serum endotoxin and both IgG and IgM antiendotoxin core antibodies (EndoCAbs) were measured on admission. Attacks were classified as mild (n = 51) or severe (n = 21) according to the criteria of the Atlanta International Symposium of 1992. RESULTS: Hypocalcemia was significantly more frequent (86% versus 39%, p < 0.001) and reached significantly lower levels during severe attacks than during mild attacks (median [interquartile range], 2.06 [1.78-2.17] mmol/L, versus 2.23 [2.15-2.30] mmol/L; p < 0.001). Endotoxemia was present in a significantly greater proportion of patients with severe disease (71% versus 41%; p = 0.037), and serum IgM and IgG EndoCAbs were significantly depleted during severe attacks in comparison with mild attacks (p = 0.007 and p = 0.039, respectively). A negative and significant correlation was observed between endotoxemia and both the admission and lowest serum concentrations of adjusted calcium (r = -0.424 and p = 0.022; r = -0.383 and p = 0.037, respectively), and the latter correlated significantly with serum IgG EndoCAb concentrations (r = 0.251; p = 0.036). CONCLUSION: Systemic endotoxin exposure appears to play a significant role in the development of hypocalcemia in patients with acute pancreatitis.

Preoperative and intraoperative predictors of postoperative morbidity, poor graft function, and early rejection in 190 patients undergoing liver transplantation.

HYPOTHESIS: Preoperative and intraoperative variables predict in part adverse outcome after liver transplantation. DESIGN: Prospective, blinded, cohort study. SETTING: Tertiary care hospital. SUBJECTS: A total of 190 adult patients undergoing primary liver transplantation. MAIN OUTCOME MEASURE: Adverse outcome was prospectively defined as either in-hospital death or prolonged postoperative hospitalization (>14 days) associated with morbidity. Potential preoperative and intraoperative risk factors were collected. Associations were tested by univariate analysis followed by multivariate analysis in which preoperative factors were entered before intraoperative factors. RESULTS: Adverse outcome occurred in 44.7% of patients. Incidences of other complications were as follows: in-hospital mortality (8.4%), primary graft nonfunction (4.2%), poor early graft function (1.1%), and early rejection (31.2%). Univariate predictors of adverse outcome were United Network for Organ Sharing status (P =.003), Child-Turcotte-Pugh score (P =.02), POSSUM physiological score (P =.002), recipient age (P =.01), preoperative serum high-density lipoprotein cholesterol level (P =.03), preoperative serum creatinine level (P =.002), preoperative serum total IgG level (P =.004), duration in hospital preoperatively (P =.03), operative duration (P<.001), allogeneic erythrocyte transfusions (P<.001), total intraoperative fluids (P =.002), and use of inotropic agents (P =.01). In the final multivariate model, predictors of adverse outcome were United Network for Organ Sharing status (P =.03), recipient age (P =.002), and total intraoperative fluids (P =.04). Most patients who died or had a prolonged hospitalization exhibited dysfunction of more than 1 organ system, including pulmonary, renal, and infectious complications. CONCLUSIONS: Adverse outcome occurs frequently after liver transplantation, usually involves multiple organ systems, and is predicted in part by several preoperative and intraoperative factors.

Endotoxin-neutralizing capacity of serum from cardiac surgical patients.

OBJECTIVE: To determine if endotoxin core antibody (EndoCAb) from the serum of cardiac surgical patients neutralizes endotoxin in an ex vivo biologic assay. DESIGN: Prospective blinded cohort study. SETTING: Academic medical center. PARTICIPANTS: Patients (n = 203) undergoing cardiac surgery. INTERVENTIONS: Sera were obtained from patients preoperatively. MEASUREMENTS AND MAIN RESULTS: EndoCAb levels were determined by enzyme-linked immunosorbent assay. Sera were incubated for 15 minutes at 37 degrees C with varying concentrations of endotoxin from a clinically relevant bacterium (Escherichia coli serotype O18), then tested for the presence of endotoxin activity using the validated Limulus amebocyte lysate assay. Median (interquartile range) IgM and IgG EndoCAb levels were 118 median units (range, 31 to 259 median units) and 208 median units (range, 108 to 401 medium units). Increasing levels of IgM EndoCAb were associated with increased neutralization of endotoxin (p < 0.0001). Increasing levels of IgG EndoCAb were associated with increased neutralization of endotoxin (p < 0.0001). An additive effect of IgM and IgG EndoCAb levels on endotoxin neutralization was observed without evidence of synergistic or plateau effects. EndoCAb levels did not completely predict serum neutralization capacity. CONCLUSION: Anti-EndoCAbs of both classes (IgM and IgG) were able to neutralize lipopolysaccharide from a clinically relevant bacterium in an ex vivo model. Neither Igm nor IgG appeared to be more capable of neutralization in this model. These antibodies did not completely predict neutralization capacity; other endogenous factors in human serum must be capable of lipopolysaccharide neutralization.

Decreased endotoxin immunity is associated with greater mortality and/or prolonged hospitalization after surgery.

BACKGROUND: Patients undergoing noncardiac surgery often develop postoperative morbidity, potentially attributable to endotoxemia and the systemic inflammatory response syndrome. Endogenous antibodies to endotoxin may confer protection from endotoxin-mediated toxicity. The authors sought to determine the association of preoperative antiendotoxin immunity and death or prolonged hospitalization in a broad population of general surgical patients undergoing major surgery. METHODS: To test the hypothesis that low preoperative serum antiendotoxin core antibody (EndoCAb) concentration is an independent predictor of adverse outcome after general surgery, 1,056 patients undergoing routine noncardiac surgery were enrolled into a prospective, blinded, cohort study. Immunoglobulin M EndoCAb, immunoglobulin G EndoCAb, total inmunoglobulin M, and immunoglobulin G concentrations were measured in serum obtained preoperatively. A physiologic risk score using the established POSSUM criteria was assigned preoperatively to each patient. The primary predefined composite end point (postoperative complication) was either in-hospital death or postoperative length of stay greater than 10 days. Multivariate logistic regression was used to test the study hypothesis. RESULTS: Overall, postoperative complication occurred in 234 of the 1,056 patients (22.1%). Lower immunoglobulin M EndoCAb concentration (P = 0.006) predicted increased risk of postoperative complication independent of POSSUM physiologic risk score (P < 0.001). In contrast, total immunoglobulin M and total immunoglobulin G concentrations did not predict adverse outcome. Complications involved multiple organ systems and were generally unrelated to the type or site of surgery, consistent with the systemic inflammatory response syndrome. CONCLUSIONS: Adverse outcome after routine noncardiac surgery is common and is predicted in part by low concentrations of EndoCAb. The authors' findings suggest that endotoxemia may be a cause of postoperative morbidity after routine noncardiac surgery.

Endotoxin, cytokines and lipid peroxides in children with intussusception.

BACKGROUND: Intussusception is a relatively common paediatric surgical emergency. The aim of this study was to investigate selected inflammatory mediators in children with acute intussusception and to identify potentially useful plasma markers of clinical outcome. METHODS: Clinical, radiographic, operative and pathological details were recorded prospectively of all children presenting to a single institution with a confirmed diagnosis of acute intussusception during 1 year. Paired acute and convalescent venous blood samples were collected in a standard manner for blinded analysis of the following: malondialdehyde, C-reactive protein (CRP), interleukin (IL) 6, neopterin, tumour necrosis factor alpha, endotoxin, and immunoglobulin (Ig) G and IgM antiendotoxin core antibody (EndoCAb). RESULTS: Thirty-two consecutive children (23 boys, nine girls) with a median age of 4 months were studied. Acute ileocolic intussusception was managed by air enema reduction (n = 19), operative reduction (n = 8) or surgical resection (n = 5). Peripheral blood cultures were sterile. Acute levels of plasma IL-6, neopterin and CRP were significantly raised in comparison to both normal laboratory ranges and convalescent samples (P < 0.001). Using stepwise discriminant analysis, CRP was identified as the best variable at distinguishing between the three treatment groups (P < 0.001). IgM EndoCAb concentrations were significantly greater in the convalescent sera of all the patients (P < 0.001). CONCLUSION: Acute ileocolic intussusception in childhood is associated with endotoxinaemia and significantly raised levels of circulating inflammatory cytokines. Plasma CRP at diagnosis showed a statistically significant positive correlation with disease severity.

Preparation and preclinical evaluation of a novel liposomal complete-core lipopolysaccharide vaccine.

Our objective is to develop a prophylactic vaccine strategy that can be evaluated for surgical and other high-risk hospitalized patients. In this paper, we describe the preparation and preclinical evaluation of a liposomal complete-core lipopolysaccharide (LPS) vaccine that is nontoxic and broadly antigenic. Complete-core (Ra-chemotype) LPSs were isolated from four gram-negative bacterial strains (Escherichia coli K-12, E. coli R1, Pseudomonas aeruginosa PAC608, and Bacteroides fragilis), mixed together to form a cocktail of complete-core LPSs, and then incorporated into multilamellar liposomes consisting of dimyristoyl phosphatidyl choline, dimyristoyl phosphatidylglycerol, and cholesterol in a 4:1:4 molar ratio. The endotoxic activities of these LPS-containing liposomes were less than 0.1% of the endotoxicities of the original free LPSs as measured by the Limulus amoebocyte lysate assay. In vivo administration of liposomal complete-core LPS mixed with Al(OH)(3) to rabbits resulted in no pyrogenicity or overt toxicity over a 7-day period. In immunoblots, sera from rabbits following active immunization elicited cross-reactive antibodies to a large panel of rough and smooth LPSs from numerous clinically relevant gram-negative bacteria, including E. coli (serotypes O1, O4, O6, O8, O12, O15, O18, O75, O86, O157, and O111), P. aeruginosa (Fisher-Devlin serotypes 1, 2, and 3, which correspond to International Antigenic Typing Scheme types 6, 11, and 2, respectively), Klebsiella pneumoniae (serotypes O1, O2ab, and O3), B. fragilis, and Bacteroides vulgatus. Active immunization of mice with liposomal complete-core LPS provided protection against a lethal challenge with E. coli O18 LPS. The vaccine tested was nontoxic, nonpyrogenic, and immunogenic against a wide variety of pathogens found in clinical settings.

Quality assurance by a commercial flow cytometry method of leucodepletion of whole blood donations: initial application of universal testing and proposals for a batch-release sampling plan.

We have used the Becton-Dickinson LeucoCOUNT test to monitor residual leucocytes in whole blood by flow cytometry following leucodepletion filtration. This test was found to be quick, robust and reliable, and allowed measurement of residual leucocytes down to 2.25 x 104 leucocytes per unit, which was found to match approximately the limit of filter proficiency. The results of testing > 1000 units showed a lognormal distribution with means between log(10) 4.864 (0.73 x 105) and log(10) 5.016 (1.04 x 105) leucocytes per unit in the three different homogeneous groups of filtered units studied. The numbers of units with residual leucocytes exceeding the 5 x 106 upper threshold were 1/577, 2/457 and 0/87 in these groups. The filtration processes were validated according to the published BEST working party guidelines and were well within the 99% confidence and 95% tolerance target for the 5 x 106 upper threshold set for the UK. A sampling plan based on British Standard BS 6001 with elimination of outliers by the extreme studentized deviate (Grubbs' test) was evaluated on random samples from these groups and has been adopted locally as a prospective batch-release criterion for release of leucodepleted blood.

Exposure to bacteroides fragilis endotoxin during cardiac surgery.

UNLABELLED: Although endotoxemia has been observed during cardiac surgery, the identity of endotoxins to which patients are exposed is unknown. We tested the hypothesis that antibodies to Bacteroides fragilis (an anaerobic gut commensal and a common pathogen) decrease during cardiac surgery, thereby reflecting systemic exposure to this type of endotoxin. Serum antiendotoxin antibody levels were measured in 55 patients during routine cardiac surgery at the following times: Preoperatively, Pre-CPB (immediately before initiation of cardiopulmonary bypass [CPB]), Pre-CPB+5 (5 min after initiation of CPB), and End (end of surgery). Antiendotoxin antibody levels were determined by using enzyme-linked immunosorbent assay. Total immunoglobulin M (IgM) levels were measured by using laser nephelometry and decreases in total IgM levels were used to control changes in antiendotoxin antibody levels attributable to hemodilution. Median (interquartile range) hemodilution corrected IgM anti-B fragilis antibody levels decreased by 12% (5%-20%) from Preoperatively to End of surgery (P < 0.001). In contrast, median hemodilution corrected anti-B fragilis antibody levels did not change significantly from Pre-CPB to Pre-CPB+5, validating the correction for hemodilution. Immunoglobulin G anti-B fragilis antibody levels and IgM and immunoglobulin G anticore antibody levels decreased similarly during surgery. Intraoperatively, levels of anti-B fragilis endotoxin antibodies decreased significantly out of proportion to hemodilution. These results suggest that cardiac surgical patients are exposed to B fragilis endotoxin. IMPLICATIONS: We prospectively measured hemodilution-corrected antiendotoxin antibody levels in 55 cardiac surgical patients. We observed significant decreases in hemodilution-corrected levels of antibody to both Bacteroides fragilis and the core of endotoxin.

Lack of correlation between failure of gut barrier function and septic complications after major upper gastrointestinal surgery.

OBJECTIVE: To determine the influence of abnormal gut barrier function on the risk of septic complications in patients undergoing major resectional surgery for upper gastrointestinal cancer. SUMMARY BACKGROUND DATA: A failure of the gut mucosal barrier to exclude bacteria and endotoxin from the portal and systemic circulation is incriminated in the development of sepsis and multiple organ failure. Although the experimental data is compelling, corroborative evidence from studies in humans is sparse. This study attempted to correlate both preoperative gut barrier dysfunction and the pattern of change after surgery with septic outcome. METHODS: Sixty-eight patients undergoing curative resectional surgery for upper gastrointestinal cancer were monitored for 30-day septic morbidity (intraabdominal abscesses/empyema and pneumonia). Intestinal permeability, serum IgM and IgG anti-endotoxin antibodies (EndoCAb), and serum C-reactive protein were measured before surgery and on postoperative days 1 and 7. RESULTS: Increased intestinal permeability before surgery did not predict septic outcome. Major surgery was associated with increased intestinal permeability and evidence of endotoxin exposure. Comparing sepsis and nonsepsis groups, however, there was no significant difference in intestinal permeability, endotoxin exposure, and the acute phase response after surgery. CONCLUSIONS: This study demonstrates that gut barrier dysfunction occurs after surgery, but the magnitude of change does not differentiate patients in whom sepsis develops and those in whom it does not. Preoperative increased intestinal permeability had no predictive value for sepsis. This study failed to support the thesis that gut barrier dysfunction is directly linked to sepsis.

Distribution of cell-associated prion protein in normal adult blood determined by flow cytometry.

Leucocyte subpopulations from normally healthy individuals were identified by recognized combinations of fluorochrome-conjugated antibodies to CD markers and stained by different monoclonal antibodies (MAb) to normal cellular prion protein (PrPC), including the 3F4 MAb. Cell preparations were examined by three-colour flow cytometry. All mononuclear leucocyte subpopulations and platelets expressed PrPC, but polymorphonuclear leucocytes and red blood cells expressed little or no PrPC. The amounts of PrPC expressed by the different cells were calculated by comparison to bead standards. Mononuclear leucocytes expressed 3000-4000 molecules of antibody-reactive PrPC per cell. Resting platelets expressed around 1400 molecules of PrPC per cell, whereas activated platelets expressed around 4800 molecules of PrPC per cell. Extrapolation of these values to the amounts of the various cells in whole blood showed that platelet PrPC accounted for at least 96% of cell-expressed PrPC in blood. The PrPC on mononuclear cells and platelets was sensitive to enzymatic treatment of cells by proteinase k and phosphatidylinositol-specific phospholipase C. Certain anti-PrPC MAbs which showed equivalent intensity of staining to MAb 3F4 on fresh cells showed relative reductions of staining compared to MAb 3F4 on stored cells, indicating possible structural alterations of PrPC under these conditions.

Early increase in intestinal permeability in patients with severe acute pancreatitis: correlation with endotoxemia, organ failure, and mortality.

Sepsis accounts for 80% of deaths from acute pancreatitis. This study aimed to investigate early changes in intestinal permeability in patients with acute pancreatitis, and to correlate these changes with subsequent disease severity and endotoxemia. The renal excretion of enterally administered polyethylene glycol (PEG) 3350 and PEG 400 was measured within 72 hours of onset of acute pancreatitis to determine intestinal permeability. Severity was assessed on the basis of APACHE II scores and C-reactive protein measurements. Serum endotoxin and antiendotoxin antibodies were measured on admission. Eight-five patients with acute pancreatitis (mild in 56, severe in 29) and 25 healthy control subjects were studied. Urinary excretion of PEG 3350 (median) was significantly greater in patients who had severe attacks (0.61%) compared to those with mild disease (0.09%) and health control subjects (0.12%) (P <0. 0001), as was the permeability index (PEG 3350/400 excretion) (P <0. 00001). The permeability index was significantly greater in patients who subsequently developed multiple organ system failure and/or died compared with other severe cases (0.16 vs. 0.04) (P = 0.0005). The excretion of PEG 3350 correlated strongly with endotoxemia (r = 0.8; P = 0.002). Early increased intestinal permeability may play an important role in the pathophysiology of severe acute pancreatitis. Therapies that aim to restore intestinal barrier function may improve outcome.