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PubMed2000

Interferon-gamma-primed monocytoid cell lines: optimizing their use for in vitro detection of bacterial pyrogens.

In order to reduce animal testing for quality control of pharmaceutical agents intended for parenteral use, the Limulus amebocyte lysate (LAL) assay is now being accepted in many cases as an alternative to measuring pyrogenic activity of samples in rabbits. However, since the LAL test is specific for cell wall components from Gram-negative bacteria and is sometimes difficult to perform in samples containing large amounts of protein, this alternative still leaves a considerable diagnostic gap. Here, we have optimized a previously established test based on assessing the formation of neopterin or nitrite in interferon-gamma-treated human (THP-1) or murine (J774A.1, RAW264.7) monocytoid cell lines, respectively, in response to bacterial pyrogens. Optimal results were obtained either with THP-1 cells in serum-containing media and using a high concentration of interferon-gamma (IFN-gamma) or with RAW264.7 cells in serum-free media and independent of the IFN-gamma dose. Results were significantly correlated with those obtained by another cell-culture-based assay in which formation of tumor necrosis factor-alpha by THP-1 1G3 cells was assessed. Also in RAW264.7 murine monocytoid cells, formation of nitrite and of tumor necrosis factor-alpha in response to a variety of samples was correlated. Samples shown to be pyrogenic in rabbits in a previous study were unambiguously detected with the test presented here. As expected, the LAL test was negative with cell-free supernatants from Staphylococcus aureus66 kDa). Taken together, these results indicate that the use of monocytoid cell lines and the detection of metabolites which are triggered in the course of immunostimulation could fill the gap left by the LAL test and help to further reduce animal testing for pyrogens.

Animal Testing Alternatives↗
PubMed1980

A brief report of gram-negative bacterial endotoxin levels in airborne and settled dusts in animal confinement buildings.

Gram-negative bacterial endotoxins, implicated in adverse worker health responses, were found in settled and airborne dust samples obtained from poultry and swine confinement units. Results of the Limulus amebocyte lysate gel test found endotoxin levels in dust samples ranged from 4.5 to 47.7 micrograms of FDA Klebsiella endotoxin equivalents/gm. Differences in endotoxin levels between dust samples may have been due to variables in time, geographic locations, confined animals, confinement buildings and equipment, and methods of sample collection. Animal confinement workers are potentially exposed to large amounts of gram-negative bacterial endotoxins; however, the respiratory health effects of such exposures to animal confinement workers have yet to be determined.

Agricultural Workers' Diseases↗
PubMed2004

Inflammatory activity in river-water samples.

Contamination of the urban aquatic environment with chemical and biological substances could have a long-term impact on human health because these substances threaten the integrity of the urban ecosystem and the availability of high-quality water for recreation and consumption. In light of this, the aim of the present study was to assess the potential immunological effects of water sampled at various sites along the River Saale near the city of Halle (in the state of Sachsen-Anhalt, Germany). For the control, Ficoll-separated peripheral blood mononuclear cells (PBMC) of healthy donors were cultured for 24 h in either filter-sterilized river water or drinking-water samples. Cell vitality was assessed using the MTT bioassay. Cytokines in culture supernatants were measured by ELISA. Endotoxin concentrations in the water samples were assessed by the limulus amoebocyte lysate (LAL) test. River water and drinking water showed comparably weak cytotoxic effects on PBMC. Drinking water did not exert any effect on cytokine secretion. In contrast, all river-water samples triggered secretion of proinflammatory cytokines, as shown for TNF-alpha, IL-1beta, and IL-6. Free endotoxin was detected in all river-water samples. However, the highest inflammatory activity regarding induction of all three cytokines, as well as the highest endotoxin content as determined by LAL, was found in a water sample taken immediately downstream of a wastewater treatment plant. Inhibition studies using the monoclonal anti-CD14 antibody biG14, which is known to suppress binding of lipopolysaccharide (LPS) to CD14 via binding CD14 itself, revealed that free endotoxin was indeed the major inducer of proinflammatory cytokines in the river-water samples. Taken together, the results suggest that the microorganism-derived endotoxin is a widely distributed contaminant in the urban aquatic environment that should be considered in routine monitoring and in assessing ecosystem and human health.

Antibodies, Monoclonal↗
PubMed1981

Studies on Escherichia coli vaccines: estimation of endotoxins in veterinary vaccines.

Fifteen batches of E. coli vaccines for pigs, from three different manufacturers, were subjected to a quantitative form of the in vitro Limulus-amoebocyte-lysate (LAL) test for their free endotoxin content. A batch of vaccine associated with abortions in pregnant sows was found to contain a much higher level of free endotoxin than batches of vaccine not associated with abortion. The evidence of these assays suggests that they will be useful in the quality control of E. coli vaccines.

Abortion, Veterinary↗
PubMed1982

Endotoxaemia in Vibrio El Tor cholera.

Endotoxaemia and endotoxin-induced changes were sought in Nigerian patients presenting with cholera/diarrhoea. The organism was Vibrio cholerae, bio-type El Tor, serotype Hikojima. The limulus amoebocyte lysate gelation test was used qualitatively by the clot method, whilst a spectrophotometric method was used quantitatively to measure endotoxin levels. 25 acutely ill patients tested had detectable endotoxaemia by the Escherichia coli endotoxin standard. The highest endotoxin level was found in a patient with sub-conjunctival haemorrhage. Changes in platelet counts, the detection of complement breakdown product C3d in plasma, the elevation of fibrin degradation products, the finding of elevated, normal or depressed C3 levels and the absence of circulating immune complexes, suggest a pathogenic role for endotoxin in Vibrio cholerae El Tor diarrhoea.

Acute Disease↗
PubMed1989

The occurrence of endotoxin in sows with coliform mastitis.

The purpose of this study was to determine the role of endotoxins in the etiology of coliform mastitis /CM/ in sows under field conditions, by using the Limulus amebocyte lysate /LAL/ test for the detection of endotoxins in the blood of pigs. For this purpose, blood samples from 40 healthy sows and 46 sows with clinical signs of CM were drawn once between 24 and 72 h post partum and tested accordingly. Only one clinically healthy sow 2.5% showed the presence of endotoxins in the serum. In sows clinically affected with one or more symptoms of the CM, the bacterial toxins were detected in 15 cases /32.5%/. The results of these studies support the observation that in only a certain percentage of all CM cases the occurrence of endotoxin in blood can be shown. Sampling time in relation to the occurrence of clinical symptoms and sampling frequency may have had an influence on the detectability of circulating endotoxins. Consequently, not all cases with endotoxemia may have been identified. The usefulness of the LAL test for endotoxin detection in the blood of pigs was confirmed.

Journal Article↗
PubMed1991

Serum levels of tumor necrosis factor determine the fatal or non-fatal course of endotoxic shock.

The role of tumor necrosis factor alpha (TNF alpha) in endotoxin-induced shock was investigated in pigs receiving 5 micrograms kg-1 of Escherichia coli endotoxin (LPS) during 60 min of continuous infusion into the superior mesenteric artery. LPS concentration in aortic plasma, as determined by a chromogenic Limulus amoebocyte lysate (LAL) test, reached a peak of approximately 1000 ng l-1 during LPS infusion, and declined rapidly after discontinuation of the infusion. Serum TNF levels were determined by a bioassay using the L929 murine transformed fibroblast line. Eight of the 17 animals infused with LPS died within 30 min after beginning LPS administration, while the other 9 pigs survived beyond the experimental observation period of 3 h, although they were in a state of shock. No difference in LPS concentration was found between the survivors and the non-survivors. However, the serum TNF levels in non-survivors were significantly higher than in survivors when measured at 30 min after beginning LPS administration. In survivors, the peak increase in serum TNF levels was measured at 60 min after the beginning of LPS injection and returned rapidly to the baseline values. Although the role of TNF inducing rapid death seems to be dominant, the hemodynamic, hematology and blood chemistry disturbances seen during shock continued in survivors long after the return of TNF to baseline levels. These findings indicate that besides TNF other mediators are also involved in the LPS infusion-induced shock.

Anaphylaxis↗
PubMed1995

Quality of hemodialysis water: a 7-year multicenter study.

Since dialysis was introduced 30 years ago, constant progress in technology permitted shortening the length of hemodialysis (HD) sessions. Through growing concerns about the inadequacy of tap water for dialysate production, hospitals soon opted for water treatment systems dedicated to HD. Nonetheless, persistent bacterial contamination and the occurrence of pyrogenic reactions were reported in some HD centers. Several factors contributing to this situation were identified. After the introduction of highly permeable synthetic membranes in the late 1970s, microbiologic problems reappeared. Thus, in 1977, the Centers for Disease Control and Prevention (CDC) issued proposed guidelines for HD water quality, followed in 1981 by an American National Standard for HD water, issued by the Association for the Advancement of Medical Instrumentation (AAMI). This Standard was also followed in Canada up to 1986, at which time a National Standard for Canada was released by the Canadian Standards Association (CSA). This prompted the Laboratoire de santé publique du Québec (LSPQ) to implement in the Province of Québec a voluntary HD water quality monitoring program. All 36 HD centers in the Province agreed to participate. The program was launched in February 1987. Water was sampled monthly for bacteria over a 7-year period (February 1987 to January 1994), and every 3 months for pyrogen and chemicals. Participation was more than 95%. Bacteriologic samples were processed in duplicate on heterotrophic plate count agar by the pour plate technique. Incubation was for 48 +/- 3 hours at 35 +/- 0.5 degrees C, and the colonies were counted on a Quebec colony counter (New Brunswick Scientific Co, New Brunswick, NJ). Pyrogen determinations were made using the limulus amebocyte lysate (LAL) test on 1:20 sample dilution by the gel-clot method. Chemical elements were measured by inductively coupled plasma emission, graphite furnace absorption, conductivity, ultraviolet light absorption, or colorimetry. Only fully treated HD water samples were selected from the 11,000 water samples received. Of the 5,820 samples retained for this study, 3,547 were for bacterial, 1,112 for pyrogen, and 1,161 for chemical analyses. Overall compliance to the CSA Standard was 70% for bacteria, 56% for pyrogen, and 86% for chemistry. The performance of different types of water treatments were compared and discussed; the best overall compliance was obtained by reverse osmosis combined with deionization (RO + DI). The type of water treatment that proved most popular was RO alone, which was used by 22 HD centers (61%).(ABSTRACT TRUNCATED AT 400 WORDS)

Bacteria↗
PubMed1997

Plasma cytokines and endotoxin levels in patients with severe injury and their relationship with organ damage.

In 17 patients plasma TNF-alpha and IL-8 were assayed with enzyme-linked immunosorbent assay. IL-6 activity in plasma was determined by bioassay with IL-6-dependent cell line 7TD1. The limulus amoebocyte lysate chromogenic test was used for plasma endotoxin assay. Plasma cytokine levels in injured patients were significantly increased. Plasma TNF-alpha was shown to be increased earlier, while an increase in plasma IL-6 and IL-8 levels occurred late, all of which were shown to be significantly positively correlated with ISS, cardiac and hepatic enzyme activities, and index of renal function. In addition, obvious endotoxaemia occurred at an early stage of injuries, which was respectively significantly correlated with ISS and plasma TNF-alpha, IL-6 and IL-8 levels. Severe injuries could induce increased successive release of TNF-alpha, IL-6 and IL-8, and obvious endotoxaemia. The post injury release of cytokines might be related to endotoxaemia, and may play an important role in the development of organ damage after injury.

Adolescent↗
PubMed1977

Activities of lectins and their immobilized derivatives in detergent solutions. Implications on the use of lectin affinity chromatography for the purification of membrane glycoproteins.

The effects of several commonly used detergents on the saccharide-binding activities of lectins were investigated using lectin-mediated agglutination of formalin-fixed erythrocytes and affinity chromatography of glycoproteins on columns of lectins immobilized on polyacrylic hydrazide-Sepharose. In the hemagglutination assays, Ricinus communis I (RCA1) and II (RCAII), concanavalin A (Con A), and the agglutinins from peanut (PNA), soybean (SBA), wheat germ (WGA), and Limulus polyphemus (LPA) were tested with several concentrations of switterionic, cationic, anionic, and nonionic detergents. It was found that increasing detergent concentrations eventually affected hemagglutination titers in both test and control samples, and the highest detergent concentrations not affecting lectin hemagglutinating activities were determined. The effects of detergents on specific binding of [3H]fetuin and asialo[3H]fetuin to and elution from columns of immobilized lectins were less severe when compared with lectins in solution, suggesting that the lectins are stabilized by covalent attachment to agarose beads. Nonionic detergents did not affect the binding efficiency of the immobilized lectins tested at concentrations used for membrane solubilization while cationic and zwitterionic detergents caused significant inhibition of Con A- and SBA-Sepharose activities. In sodium deoxycholate (greater than 1%) only RCAI-Sepharose retained its activity, whereas the activities of the other lectins were reduced dramatically. Low concentrations of sodium dodecyl sulfate (0.05%) inhibited only the activity of immobilized SBA, but at higher concentration (0.1%) and prolonged periods of incubation (16 h, 23 degrees C) most of the lectins were inactivated. These data are compared with previous reports on the use of detergents in lectin affinity chromatography, and the conditions for the optimal use of detergents are detailed.

Chromatography, Affinity↗
PubMed2003

The effect of storage on allergen and microbial agent levels in frozen house dust.

BACKGROUND: House dust samples collected for exposure studies are often stored for variable time periods until analysis. However, there is currently no information on the effects of dust storage on the content of biocontaminants. Therefore, associations were analysed between the levels of mite allergens (Der p 1, Der f 1), cat allergen (Fel d 1) and microbial components (endotoxin, beta(1-->3)-glucan) on the one hand and the storage duration of dust samples at -20 degrees C on the other hand. METHODS: Within the framework of a study on the influences of INdoor factors and Genetics on Asthma (INGA), dust samples were collected from living room floors between June 1995 and August 1998 and extracted according to a standardized protocol. The concentrations of Der p 1, Der f 1, Fel d 1 and beta(1-->3)-glucan were determined with specific enzyme immunoassays. Endotoxin content was quantified using a chromogenic kinetic Limulus amoebocyte lysate (LAL) test. All concentrations were expressed per gram of dust RESULTS: Dust samples (n = 1236) were obtained from 655 homes in Hamburg, Hettstedt, Zerbst and Bitterfeld. Storage duration (range 8-298 days) was grouped into four categories ( 120 d). After adjustment for city of residence and season of dust sampling, means ratios comparing categories 2-4 to the first category were not statistically significant for Der p 1, Der f 1, endotoxin and beta(1-->3 glucan). However, Fel d 1 concentrations significantly declined with increased storage times of dust samples. CONCLUSIONS: Storage of house dust at -20 degrees C for up to 10 months has no effect on mite allergen, endotoxin and beta(1-->3)-glucan levels. A potential loss of Fel d 1 during storage of frozen dust samples needs further investigations by repeated measurements of allergen in identical dust samples.

Air Pollution, Indoor↗
PubMed2004

Endotoxin in storage medium of human corneal grafts and clinical course after penetrating normal-risk keratoplasty.

PURPOSE: It is well known that endotoxins in storage medium may stimulate cytokine production and expression of adhesion molecules as well as endothelial damage in human corneal grafts. It has been supposed that endotoxin exposure of corneal grafts may, therefore, cause immune reactions and lead to reduced endothelial cell count after penetrating keratoplasty. It was the purpose of this prospective study to evaluate if this hypothesis is true. METHODS: A consecutive series of 274 samples of sterile organ culture storage medium from 274 human corneal grafts was collected between August 1998 and February 1999 and tested for endotoxin using Limulus amebocyte-lysate assay (LAL) after 7 days of organ culture. Threshold endotoxin level was set at 1.0 U/ml. A total of 161 grafts were transplanted and 113 were discarded. Within the 161 corneas transplanted, 62 were grafted to normal-risk patients and 99 to high-risk patients. Only normal-risk keratoplasty patients were included in the study and followed for at least 10 months. Immune reactions, graft failures, and postoperative endothelial cell counts were recorded. RESULTS: The mean endotoxin level in organ culture medium of all transplanted grafts was 1.07+/-2.96. Mean endotoxin level in organ culture medium of discarded grafts was 1.68+/-5.76, with 71 samples being below and 42 above the threshold of 1.0 U/ml called endotoxin-negative and endotoxin-positive, respectively. In all 36 culture medium samples from the 62 grafts transplanted to the group of normal-risk keratoplasty patients were endotoxin-negative and 26 endotoxin-positive. An influence of endotoxin levels on incidence of immune reactions, graft failure, and postoperative endothelial cell counts could not be revealed in patients with normal-risk keratoplasty. CONCLUSION: Low endotoxin levels in storage medium neither seem to promote immune reactions nor to contribute to postoperative chronic endothelial cell loss in normal-risk keratoplasty patients.

Adolescent↗
PubMed2006

Biocompatibility of an enzyme-based, electrochemical glucose sensor for short-term implantation in the subcutis.

BACKGROUND: Continuous glucose measurements provide improved glycemic control and may prevent hypoglycemia and long-term complications of diabetes. One of the most promising techniques is the short-term implantation of electrochemical glucose sensors in subcutis. However, the inflammatory reaction to these sensors may lead to bioinstability of sensor measurements. The purpose of the present investigation was to examine factors contributing to the observed subcutaneous inflammatory reaction to an enzyme-based electrochemical glucose sensor for continuous glucose measurements. The sensor biocompatibility was assessed in vitro and in vivo. METHODS: A toxicological assessment was performed on sensor materials and leachables, and the endotoxin content of sensors was determined by a Limulus amoebocyte lysate (LAL) test. Moreover, as a consequence of permanent penetration of the skin by the sensor the role of bacterial migration to the tissue was investigated. In vivo biocompatibility was investigated through histological examination of implanted sensor membranes for 3 days in pigs. Additionally, the effect of needle size and type (normal vs. inserter needle) on tissue trauma at sensor insertion was evaluated, and the healing of subcutis was assessed histologically from 3 to 14 days after removal of sensors. RESULTS: The toxicological assessment and the LAL test showed no concerns in a 3-day implantation scenario, and bacterial migration to the subcutis could not be detected. The histological examination showed that a reduction in needle size reduced the extent of inflammation to very low levels, and that the different sensor membranes showed similar extent and type of inflammation. Additionally, the extent of subcutaneous tissue reaction after removal of sensors declined gradually over time and returned to near-normal levels after 2 weeks. CONCLUSION: The electrochemical enzyme-based glucose sensor for continuous glucose measurements in subcutis is acceptable from a biocompatibility point of view. Reducing the inserter needle in size reduces the trauma induced at sensor implantation to neglible levels. Furthermore, the tissue reaction to the sensor returns to near-normal 2 weeks after the sensor has been removed following a 3-day implantation period.

Animals↗
PubMed1998

Synthesis and characterization of lipooligosaccharide-based conjugates as vaccine candidates for Moraxella (Branhamella) catarrhalis.

Moraxella (Branhamella) catarrhalis is an important cause of otitis media and sinusitis in children and of lower respiratory tract infections in adults. Lipooligosaccharide (LOS) is a major surface antigen of the bacterium and elicits bactericidal antibodies. Treatment of the LOS from strain ATCC 25238 with anhydrous hydrazine reduced its toxicity 20,000-fold, as assayed in the Limulus amebocyte lysate (LAL) test. The detoxified LOS (dLOS) was coupled to tetanus toxoid (TT) or high-molecular-weight proteins (HMP) from nontypeable Haemophilus influenzae through a linker of adipic acid dihydrazide to form dLOS-TT or dLOS-HMP. The molar ratios of dLOS to TT and HMP conjugates were 19:1 and 31:1, respectively. The antigenicity of the two conjugates was similar to that of the LOS, as determined by double immunodiffusion. Subcutaneous or intramuscular injection of both conjugates elicited a 50- to 100-fold rise in the geometric mean of immunoglobulin G (IgG) to the homologous LOS in mice after three injections and a 350- to 700-fold rise of anti-LOS IgG in rabbits after two injections. The immunogenicity of the conjugate was enhanced by formulation with monophosphoryl lipid A plus trehalose dimycolate. In rabbits, conjugate-induced antisera had complement-mediated bactericidal activity against the homologous strain and heterologous strains of M. catarrhalis. These results indicate that a detoxified LOS-protein conjugate is a candidate for immunization against M. catarrhalis diseases.

Animals↗
PubMed2002

Structural decomposition and heterogeneity of commercial lipoteichoic Acid preparations.

Fractionation of commercial preparations of lipoteichoic acids (LTA) by hydrophobic interaction chromatography (HIC) and nuclear magnetic resonance spectroscopy revealed very inhomogeneous compositions and decomposition of the LTA structure: LTA content of the preparations averaged 61% for Streptococcus pyogenes, 16% for Bacillus subtilis, and 75% for Staphylococcus aureus. The decomposition was characterized by a loss of glycerophosphate units as well as alanine and N-acetylglucosamine substituents. All preparations contained-to varying degrees-non-LTA, non-lipopolysaccharide (LPS) immunostimulatory components as indicated by their elution profile in HIC, lack of phosphate, and negative Limulus amoebocyte lysate (LAL) test results. After purification, the commercial LTA from Bacillus subtilis and S. pyogenes but not LTA from S. aureus induced the release of tumor necrosis factor alpha, interleukin 1 beta (IL-1beta), IL-6, and IL-10 in human blood. While pure LTA are negative in the LAL assay, endotoxin equivalents of more than 10 ng of LPS/mg of LTA were found in the commercial preparations. Taken together, these data indicate that these crude preparations with relatively high endotoxin contamination are not suitable for characterizing the activation of immune cells by LTA.

Bacillus subtilis↗
PubMed1990

Lactulose inhibits endotoxin induced tumour necrosis factor production by monocytes. An in vitro study.

Preoperative oral treatment with lactulose is used to prevent complications after surgery in patients with obstructive jaundice. The effect is perhaps the result of an inactivation of gut derived endotoxins but the exact mechanism of action is, however, unknown. Tumour necrosis factor is an important mediator of endotoxin toxicity. The cytokine tumour necrosis factor is mainly produced by mononuclear phagocytes. In this study, the effect of lactulose on the endotoxin induced tumour necrosis factor release by monocytes was investigated. The direct effect of lactulose on endotoxin was tested in a chromogenic limulus amoebocyte lysate assay. Polymyxin B a known inactivator of endotoxin was used as control in both experiments. Lactulose has a limited capacity to inactivate endotoxin as measured in the endotoxin assay. In contrast lactulose significantly reduced endotoxin induced tumour necrosis factor production by monocytes. In conclusion lactulose inhibits tumour necrosis factor production by a direct inhibitory effect on monocytes, rather than by inactivation of endotoxin. Because tumour necrosis factor is an important mediator of endotoxin toxicity, this inhibitory effect could explain the beneficial effect of lactulose in obstructive jaundice.

Cells, Cultured↗
PubMed1988

[The effect of water rinsing in removal of endotoxin from exposed root].

Previous studies have revealed that periodontally involved exposed roots show evidence of biologic toxicity possibly due to endotoxin from subgingival micro-organisms. Eleven periodontally involved teeth and five periodontally healthy teeth comprised the material. The teeth were rinsed in ultrasonic cleaner containing pyrogen-free water for one hour. The samples were taken from the solution after 1, 5, or 60 minutes. With the aim of removing residual endotoxin on the roots not eliminated by rinsing, each tooth was subjected to extraction of endotoxin with 45% phenol in water for 90 minutes at 65 degrees C. After extraction of endotoxin, the root surface of the involved teeth was further planned with a hand scaler. Endotoxin was extracted from the particles removed from the root surfaces. All samples were tested for endotoxin by limulus amoebocyte lysate assay. The amount of endotoxin from involved teeth was 4,500 +/- 2,000 ng per tooth and from healthy teeth it was 370 +/- 120 ng per tooth. The amount of residual endotoxin on roots from involved teeth was 32 +/- 20 ng per tooth and from healthy teeth it was 1.2 +/- 1.1 ng per tooth. The amount of endotoxin collected from particles removed from previously rinsed involved root surfaces was 1.5 ng per tooth. The rate of removal of endotoxin by water rinsing of a tooth was 64 +/- 25% in 1 minute, 85 +/- 16% in 5 minutes, 99 +/- 0.6% in 60 minutes.

Endotoxins↗
PubMed1993

Endotoxin liberation from Neisseria meningitidis correlates to their ability to induce procoagulant and fibrinolytic factors in human monocytes.

Endotoxin released from different strains of Neisseria meningitidis were studied for their ability to induce procoagulant (tissue factor, TF), fibrinolytic (plasminogen activator, PA) and antifibrinolytic (plasminogen activator inhibitor 2, PAI-2) factors in human monocytes. Two meningococcal strains that liberate endotoxin (E+; 270+ and 840+) and 2 non-liberating (E-; 270- and 840-) strains were used. The endotoxin activity in culture filtrates of these strains was monitored with the Limulus amoebocyte lysate (LAL) test. There was a marked difference between E+ and E- strains in their ability to liberate endotoxin. Suspensions of whole bacteria of all 4 strains induced a significant (14-19-fold) increase in monocyte TF expression when present in concentrations > 10(5) CFU/ml. At lower concentrations (10(4) CFU/ml), E+ strains were clearly more potent stimulators of TF synthesis than E- strains. Culture filtrates of E+ strains were up to 10(4)-fold more potent in inducing TF synthesis than filtrates from E- strains. This marked difference in inducing potency between E+ and E- strains was also observed when monocyte PAI-2 synthesis was examined. The PA expression, on the other hand, was suppressed when monocytes were incubated in the presence of culture filtrates, especially filtrates from the E+ strains. The increased procoagulant and antifibrinolytic activity, together with reduced profibrinolytic activity of monocytes, was closely correlated to the amount of endotoxin measured in the culture filtrates. These changes may contribute substantially to the coagulopathic state seen during systemic meningococcal disease.

Cells, Cultured↗