Bacterial sensing, cell signaling, and modulation of the immune response during sepsis

ABSTRACT: Since the definition of systemic inflammatory response syndrome/sepsis was originally proposed, a large amount of new information has been generated showing a much more complex scenario of inflammatory and counterinflammatory responses during sepsis. Moreover, some fundamental mechanisms of sensing and destroying invading microorganisms have been uncovered, which include the discovery of TLR4 as the lipopolysaccharide (LPS) gene, implications of innate immune cells as drivers of the adaptive response to infection, and the modulation of multiple accessory molecules that stimulate or inhibit monocyte/macrophage and lymphocyte interactions. The complexity of the infection/injury-induced immune response could be better appreciated with the application of genomics and proteomics studies, and LPS was a useful tool in many of these studies. In this review, we discuss aspects of bacterial recognition and induced cellular activation during sepsis. Because of the relevance of endotoxin (LPS) research in the field, we focus on LPS and host interactions as a clue to understand microorganisms sensing and cell signaling, then we discuss how this response is modulated in septic patients.     

Lipopolysaccharide sensing an important factor in the innate immune response to Gram-negative bacterial infections: benefits and hazards of LPS hypersensitivity

In this review, we summarize our investigations concerning the differential importance of CD14 and LBP in toll-like receptor 4 (TLR4)/myeloid differentiation protein-2 (MD-2)-mediated signaling by smooth and rough-form lipopolysaccharide (LPS) chemotypes and include the results obtained in studies with murine and human TLR4-transgenic mice. Furthermore, we present more recent data on the mechanisms involved in the induction of LPS hypersensitivity by bacterial and viral infections and on the reactivity of the hypersensitive host to non-LPS microbial ligands and endogenous mediators. Finally, the effects of pre-existing hypersensitivity on the course and outcome of a super-infection with Salmonella typhimurium or Listeria monocytogenes are summarized.     

Rifampicin-associated segmental necrotizing glomerulonephritis in staphylococcal endocarditis

Segmental necrotising glomerulonephritis has been reported as complication of rifampicin therapy in patients receiving treatment for tuberculosis. Changing epidemiology of infections such as infective endocarditis (IE) has led to an increase in the use of rifampicin for Staphylococcal infections. We describe a case of a patient with Staphylococcal IE who developed acute renal failure secondary to a segmental necrotising glomerulonephritis while being treated with rifampicin, and review the literature regarding this complication of rifampicin therapy.     

Oligosaccharides of Hyaluronan activate dendritic cells via toll-like receptor 4.

Low molecular weight fragmentation products of the polysaccharide of Hyaluronic acid (sHA) produced during inflammation have been shown to be potent activators of immunocompetent cells such as dendritic cells (DCs) and macrophages. Here we report that sHA induces maturation of DCs via the Toll-like receptor (TLR)-4, a receptor complex associated with innate immunity and host defense against bacterial infection. Bone marrow-derived DCs from C3H/HeJ and C57BL/10ScCr mice carrying mutant TLR-4 alleles were nonresponsive to sHA-induced phenotypic and functional maturation. Conversely, DCs from TLR-2-deficient mice were still susceptible to sHA. In accordance, addition of an anti-TLR-4 mAb to human monocyte-derived DCs blocked sHA-induced tumor necrosis factor alpha production. Western blot analysis revealed that sHA treatment resulted in distinct phosphorylation of p38/p42/44 MAP-kinases and nuclear translocation of nuclear factor (NF)-kappa B, all components of the TLR-4 signaling pathway. Blockade of this pathway by specific inhibitors completely abrogated the sHA-induced DC maturation. Finally, intravenous injection of sHA-induced DC emigration from the skin and their phenotypic and functional maturation in the spleen, again depending on the expression of TLR-4. In conclusion, this is the first report that polysaccharide degradation products of the extracellular matrix produced during inflammation might serve as an endogenous ligand for the TLR-4 complex on DCs.     

Excretion of radioactivity in faeces and urine of rats injected with 3H,14C-lipopolysaccharide.

The route of excretion of lipopolysaccharide (LPS) and its possible degradation in vivo was studied in rats using biosynthetically radiolabelled LPS from Salmonella abortus equi, carrying 3H activity exclusively in fatty acids and 14C activity in fatty acids and sugars. Following intravenous injection of the above LPS in AS2 rats with or without anaesthesia, excretion of radioactivity occurred mainly in the faeces and to smaller extent in urine. The rate of excretion was slow, a large part of the radioactivity being still present in the liver after 14 days. In faeces the percent recovery of 3H (18%) was lower than that of 14C (32%) suggesting loss of tritium activity and thereby of fatty acids from the excreted LPS. A similar loss of tritium was also found in the material remaining in the liver and spleen 14 days after LPS administration. In urine the material recovered during 14 days (about 7% of injected) was different from the original LPS, 70% of 14C activity being dialysable and practically all 3H activity being volatile. Similar results were also obtained following administration of the LPS intraperitoneally under anaesthesia. However, when the LPS was administered intraperitoneally without anaesthesia, in the majority of the animals, 90% of 14C and 54% of 3H was excreted in faeces within 3 days, suggesting that both route of administration and use of anaesthesia during injection influence the subsequent rate of excretion of LPS.     

Distribution and localization of endotoxin in the reticulo-endothelial system (RES) and in the main vessels of the rat during shock

The fate of endotoxin was followed with immunohistochemistry and radio-labelled lipopolysaccharide (LPS) in organs of the reticulo-endothelial system (RES), in the great vessels and the thoracic duct of rats during a 14 day period after the injection of a shock-inducing amount of endotoxin. The immunohistochemical detectability of LPS in most tissues increased continuously during the first 48 hours, showing the strongest LPS staining in the liver and adrenal gland. Macrophages were found to be the most important cells of primary LPS uptake in all organs except the adrenal glands, where endotoxin was mainly present in phagocytic vacuoles of the cortical epithelium. The comparison of results obtained with the immunoperoxidase method and radioactivity measurements revealed that at a later stage of the experiment the persisting LPS in liver and spleen looses its antigenic activity. The correlation between the appearance of LPS positive macrophages and histological signs of tissue injury during endotoxin shock is striking.     

Appearance of white blood cells in the cerebrospinal fluid of rats following intravenous injection of lipopolysaccharide

Intravenous injection of lipopolysaccharides (LPS), even in very low amounts (5 micrograms), leads to the appearance of white blood cells (WBC) in rat cerebrospinal fluid (CSF). The number of WBC is dependent on either dose of LPS or time of injection. The population of WBC consists of monocytes, granulocytes and lymphocytes, while red blood cells are not detectable. These findings support the pathogenetic role which LPS may play in the course of bacterial meningitis mostly in terms of pleocytosis and raised protein content in the CSF.     

Characterisation of protein co-extracted together with LPS in Escherichia coli, Salmonella minnesota and Yersinia enterocolitica

The porin proteins of Escherichia coli, Yersinia enterocolitica, and Salmonella minnesota were found to co-extract by the phenol-chloroform-petroleum ether method together with the R lipopolysaccharide of these strains. Lipopolysaccharide free protein recovered from the phenolic residue of the phenol-chloroform-petroleum ether extract migrated as a Mr 36-37,000 protein. We could demonstrate that the protein was extracted from bacteria as a high molecular weight protein-lipopolysaccharide complex. Once exposed to phenolic conditions, the protein was no longer soluble in the phenol-chloroform-petroleum ether extraction mixture, indicating a highly specific lipopolysaccharide-protein association.     

Ophthalmological manifestations of Fabry disease: a survey of patients at the Royal Melbourne Fabry Disease Treatment Centre

BACKGROUND: Fabry disease is a rare X-linked inborn error of glycosphingolipid metabolism. The aim of this study was to document the ophthalmological manifestations of patients attending the Royal Melbourne Hospital Fabry disease treatment centre. METHODS: Patients at the treatment centre had full ophthalmological examination performed. This included best corrected visual acuity, ocular motility examination and examination of the adnexae. Patients also underwent slit-lamp examination looking for the presence of features of Fabry disease, particularly examining the bulbar conjunctiva, cornea, lens and fundus. RESULTS: Thirty-four hemizygous male and 32 heterozygous female patients were recruited. The mean age of the hemizygous male patients was 37.7 years (range 18-57 years). The mean age of the heterozygous female patients was 34.6 years (range 1-78 years). Visual acuity was not affected. 97.1% of the hemizygotes and 78.1% of the heterozygotes had vascular abnormalities of the bulbar conjunctiva. Cornea verticillata was noted in 94.1% of the hemizygotes and 71.9% of the heterozygotes. 41.2% of the hemizygotes and 9.4% of the heterozygotes had anterior cataract formation. Posterior lens opacities were observed in 11.8% of the hemizygotes and none of the heterozygotes. Retinal vascular tortuosity was observed in 76.5% of the hemizygotes and 18.8% of the heterozygotes. CONCLUSION: Conjunctival vascular tortuosity was the most common manifestation in this series. Conjunctival and retinal vessel tortuosity, and corneal verticillata are frequently observed in Fabry disease. The incidence of lenticular changes is not consistently reported, but in this series and many others, it is much less common than that of corneal, conjunctival and retinal changes.