Analysis of LPS released from Salmonella abortus equi in human serum.

We investigated in which form lipopolysaccharide (LPS) is released from live bacteria incubated with human serum and whether the released LPS can interact with high density lipoprotein (HDL), the main transport protein for purified LPS in circulation. Live biotinylated Salmonella abortus equi bacteria were incubated with fresh serum (37 degrees C; 2 h). The released LPS was isolated by immunoprecipitation or immunoabsorption using specific anti-O antibodies. It was analysed and compared with purified LPS, also incubated with serum under identical conditions. Immunoprecipitation led to a 35% recovery and immunoabsorption to quantitative recovery of released or purified LPS. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and subsequent immunoblot analysis revealed that all molecular species present in the purified LPS were present in the released LPS. The rough fraction, which was co-isolated from serum together with the true smooth (O-polysaccharide-containing) molecules, exhibited S. minnesota rough mutant Rb antigenic specificity. In the immunoprecipitated material two forms of released LPS were identified. One represented LPS associated with a biotinylated bacterial component with an apparent molecular mass of 35-36 kDa, which was identified as OmpA, a major outer membrane protein. The OmpA-associated LPS was free of HDL. Another part of the released LPS was free of biotinylated bacterial components. This portion of LPS was associated with HDL, indicating that the interaction with HDL may also proceed with a part of LPS released from bacteria.     

Colchicine prevents tumor necrosis factor-induced toxicity in vivo.

Tumor necrosis factor (TNF) toxicity was induced in vivo by intravenous administration of 15 micrograms of recombinant murine TNF-alpha per kg to galactosamine-sensitized mice. Within 8 h, the animals developed a fulminant hepatitis. Intravenous administration of 0.5 mg of colchicine per kg at 19 and 4 h prior to TNF challenge protected the animals against hepatitis. Lipopolysaccharide (LPS)-stimulated, bone marrow-derived macrophages from C3H/HeN mice released significant amounts of TNF in vitro. When such macrophages were intravenously given to LPS-resistant galactosamine-sensitized C3H/HeJ mice, these animals died within 24 h. Preincubation of these transferred macrophages with colchicine did not suppress the LPS-inducible TNF release from these cells. Concordantly, administration of macrophages exposed to colchicine in vitro resulted in full lethality. However, in vivo pretreatment of C3H/HeJ mice with colchicine 19 and 4 h prior to the transfer of LPS-stimulated macrophages prevented lethality. In LPS-responsive NMRI mice which had been protected against galactosamine-LPS-induced hepatitis by pretreatment with colchicine, TNF was still released into the blood. We conclude from our findings that the in vivo protection by colchicine is mediated by blocking TNF action on target cells while the effector cells of LPS toxicity, i.e., the macrophages, remain responsive.     

A 14,000 MW lipoprotein and a glycolipid-like structure of Borrelia burgdorferi induce proliferation and immunoglobulin production in mouse B cells at high frequencies.

Sonicated preparations of Borrelia burgdorferi are able to stimulate unselected resting BALB/c spleen cells to proliferate and to produce immunoglobulin in vitro. FACS analysis of target cells prestained with an integrated cell-surface marker as well as cell-depletion experiments demonstrate that the majority of responding lymphocytes are B cells. Limiting dilution analyses of resting B cells revealed high frequencies of cells producing IgM (F 1/11-1/62) or IgG (F 1/5-1/163) in response to B. burgdorferi sonicate (B.b. sonicate). These numbers were similar to those obtained with lipopolysaccharide (LPS) (IgM: F 1/20-1/84; IgG: F 1/14-1/85) or a synthetic lipopeptide of Braun's Escherichia coli lipoprotein (IgM: F 1/15, 1/19; IgG: F 1/148, 1/34). The mitogenic structure(s) expressed by B. burgdorferi is distinct from LPS, as similar proliferative responses were obtained with B cells from LPS-resistant (C57BL/10ScCr and C3H/HeJ) and LPS-susceptible (C57BL/10ScSn, C3H/HeN) mice. Furthermore, B-cell mitogenic properties were also found in two distinct fractions of a phenol-chloroform-petroleum ether extract of B. burgdorferi: they consisted of a lipoprotein distinct from the outer surface proteins (Osp) A and B and glycolipid-like structures, respectively. These data suggest that spirochetes express a multitude of distinct structures with mitogenic activity for B cells including various lipoproteins as well as glycolipid(s).     

Longitudinal study of antibody response to lipopolysaccharides during chronic Pseudomonas aeruginosa lung infection in cystic fibrosis.

Antibodies to Pseudomonas aeruginosa from 10 cystic fibrosis patients with chronic P. aeruginosa lung infections were quantitatively and qualitatively analyzed. The development of specific antibodies in patient serum was evaluated in a longitudinal study (1972 to 1987). The concentrations and specificities of immunoglobulin G (IgG) and IgM antibodies to purified lipopolysaccharides (LPS) from clinical isolates of P. aeruginosa and to a variety of other gram-negative bacteria were studied by immunoblotting and enzyme-linked immunosorbent assay techniques. Results were compared with the number of immunoprecipitates to P. aeruginosa whole-cell extracts detected by crossed immunoelectrophoresis. IgG, but not IgM, anti-Pseudomonas LPS concentrations increased significantly at the onset of chronic infection and continued to increase during the course of the infection. There was a good positive correlation between the concentration of IgG anti-Pseudomonas LPS antibodies and the number of crossed-immunoelectrophoresis precipitins. The increases in IgG anti-LPS antibody concentrations were much higher to Pseudomonas LPS than to other LPSs. Binding studies demonstrated an increase in binding of IgG anti-Pseudomonas LPS during infection, whereas the binding of other anti-LPS antibodies decreased. Immunoblotting studies confirmed that antibodies reacted strongly with Pseudomonas LPS and weakly with Escherichia coli core-lipid A. The specificity of the reaction with Pseudomonas LPS increased with the duration of infection. It is concluded that anti-LPS response in cystic fibrosis patients during chronic P. aeruginosa infection demonstrates a marked increase in IgG anti-Pseudomonas LPS antibody concentration, specificity, and affinity. The anti-LPS enzyme-linked immunosorbent assay is proposed as a routine test to diagnose and to follow the course of chronic P. aeruginosa lung infection in patients with cystic fibrosis.     

TNF revisited: TNF-independent antitumor activity in sera of mice sensitized with Propionibacterium acnes and challenged with lipopolysaccharide

Sera of mice sensitized with bacteria and subsequently challenged with lipopolysaccharide promote hemorrhagic necrosis of tumors in vivo and display cytotoxic activity against tumor cells in vitro, which has been attributed to the induction of tumor necrosis factor (TNF). Here, we describe the induction of a previously unrecognized antitumor activity in such sera, which is distinct from TNF but displays tumor-specific cytocidal activity in vitro as well as potent tumor-regressing activity in vivo. Biochemical analysis of this activity yielded a molecular mass of approximately 150 kDa, closely resembling a novel tumoricidal factor of murine macrophages (Mphi) termed MTC 170 (Mphi tumor cytotoxin, approximate molecular mass 170 kDa), which we have previously proposed to constitute a major effector pathway for the destruction of tumor cells by activated Mphi.     

Incidence of hip fractures in Greece during a 30-year period: 1977–2007

Summary

The incidence of hip fractures doubled in Greece from 1977 to 2007 among people aged 50 and over. A mild decrease was observed after 2002, although the future trend cannot be safely anticipated at the moment. Half of all hip fractures in 2007 were derived from the age group of 80 and over.

Introduction

The purpose of this study was to determine the incidence of hip fractures during a 30-year period in Greece among people aged 50 and over and to document possible alterations in secular trends.

Methods

We studied hip fractures during 2007 and compared them with those of previous years starting from 1977 with an in-between 5-year interval (1977, 1982, 1987, 1992, 1997, 2002). Age- and sex-specific incidence was calculated, and secular trends were recorded. The relative risk of hip fracture in every age group was estimated according to the corresponding incidence of 1977.

Results

The adjusted incidence of hip fractures increased approximately 100 % throughout the study; it progressively increased from 1977 to 2002 and exhibited a mild significant decrease thereafter. The relative risk of hip fractures among subjects aged 60–69 in 2007 has declined compared with 1977 [0.85, 95 % confidence intervals (CI) 0.79–0.92, p?<?0.0005]. Among people aged 70–79, an increased relative fracture risk (1.53, 95 % CI 1.45–1.61, p?<?0.0005) was estimated in 2007 compared with 1977. People ≥80 years old were responsible for half of the hip fractures in 2007 but only for the 22.5 % of fractures in 1977. The relative fracture risk in people aged ≥80 was 2.81 times higher (95 % CI 2.64–2.98, p?<?0.0005) in 2007 than in 1977.

Conclusions

The incidence of hip fractures doubled during the last 30 years among people aged ≥50 years, although a mild decrease was observed in almost all age groups after 2002. The most affected group is 80 and over.     

Lipid A antibody titers in patients after ureterosigmoidostomy (author's transl)

Lipid A antibody titers were determined over a period of 6 months by passive hemolysis test in 10 patiens who underwent cystectomy and ureterosigmoidostomy. Preoperative the sera of all patients had no detectable anti lipid A activity. After 4 weeks 4 patients showed significant elevation of lipid A antibody titers. Only in these 4 patients the titers remained elevated up to six months. The significant elevation of lipid A antibody titers in these patients were caused by a non-obstructive infection of the kidneys. The serological determination of lipid A antibody titers appears to be an appropriate method for detecting pyelonephritis after ureterosigmoidostomy.     

Greek McGill Pain Questionnaire: validation and utility in cancer patients

In 1968, Melzack and Casey suggested that there are three major psychological dimensions of pain: sensory, affect, evaluative. These categories interact with one another to provide quantitative and qualitative information on the components of pain. In 1975, Melzack developed the McGill Pain Questionnaire, which is composed of four major parts and evaluates the qualities of pain. The aim of this study was to assess the applicability, reliability, and validity of the McGill Pain Questionnaire on a sample of Greek cancer patients receiving palliative treatment. It was administered to 114 cancer patients before the initiation of the palliative treatment, and then to 80 cancer patients during the treatment 7 days later. The results indicated that scale reliability was very good (0.95-0.97). During the pretreatment period, correlations between Present Rating Index (PRI), Present Pain Intensity (PPI), and Number of Words Chosen (NWC) ranged between 0.42 and 0.92. During the post-treatment time, the correlations ranged between 0.28 and 0.91. Only 21.8% of the words met a criterion of 30% for representativeness on the first administration of the questionnaire, and 9% met this criterion on the second. Validity was satisfactory (P < 0.005) according to "responsiveness to changes in time", as there was a statistical difference between the pretreatment and post-treatment time. Patients presented a desirable level of convergent construct validity (P < 0.05) concerning their performance status. Exploratory factor analysis was examined and two factors with eigenvalue over 1 were extracted, and they accounted for 95.2% of the variance. These results support the Greek-MPQ as a reliable and valid measure for evaluating the qualities of cancer pain in patients receiving palliative care.