Serratia marcescens internalization and replication in human bladder epithelial cells

BACKGROUND: Serratia marcescens, a frequent agent of catheterization-associated bacteriuria, strongly adheres to human bladder epithelial cells in culture. The epithelium normally provides a barrier between lumal organisms and the interstitium; the tight adhesion of bacteria to the epithelial cells can lead to internalization and subsequent lysis. However, internalisation was not shown yet for S. marcescens strains. METHODS: Elektronmicroscopy and the common gentamycin protection assay was used to assess intracellular bacteria. Via site directed mutagenesis, an hemolytic negative isogenic Serratia strain was generated to point out the importance of hemolysin production. RESULTS: We identified an important bacterial factor mediating the internalization of S. marcescens, and lysis of epithelial cells, as the secreted cytolysin ShlA. Microtubule filaments and actin filaments were shown to be involved in internalization. However, cytolysis of eukaryotic cells by ShlA was an interfering factor, and therefore hemolytic-negative mutants were used in subsequent experiments. Isogenic hemolysin-negative mutant strains were still adhesive, but were no longer cytotoxic, did not disrupt the cell culture monolayer, and were no longer internalized by HEp-2 and RT112 bladder epithelial cells under the conditions used for the wild-type strain. After wild-type S. marcescens became intracellular, the infected epithelial cells were lysed by extended vacuolation induced by ShlA. In late stages of vacuolation, highly motile S. marcescens cells were observed in the vacuoles. S. marcescens was also able to replicate in cultured HEp-2 cells, and replication was not dependent on hemolysin production. CONCLUSION: The results reported here showed that the pore-forming toxin ShlA triggers microtubule-dependent invasion and is the main factor inducing lysis of the epithelial cells to release the bacteria, and therefore plays a major role in the development of S. marcescens infections

Mycobacterium phlei cell wall complex directly induces apoptosis in human bladder cancer cells

Intact mycobacteria and mycobacterial cell wall extracts have been shown to inhibit the growth of human and murine bladder cancer. Their mechanism of action is, however, poorly understood. Mycobacterium phlei mycobacterial cell complex (MCC) is a cell wall preparation that has mycobacterial DNA in the form of short oligonucleotides complexed on the cell wall surface. In this study, we have investigated the possibility that MCC has anti-cancer activity that is mediated by two different mechanisms – a direct effect on cancer cell proliferation and viability and an indirect effect mediated by the production of interleukin 12 (IL-12), a cytokine known to possess anti-cancer activity. We have found that, although MCC is a potent inducer of IL-12 and IL-6 synthesis in monocytes and macrophages either in vitro or in vivo, it is unable to induce the synthesis of either IL-12, IL-6 or granulocyte–macrophage colony-stimulating factor (GM-CSF) by the human transitional bladder cancer cell lines HT-1197 and HT-1376. MCC is not directly cytotoxic towards these cancer cells, but induces apoptosis as determined by nuclear DNA fragmentation and by the release of nuclear mitotic apparatus protein. Mycobacterium phlei DNA associated with MCC is responsible for the induction of apoptosis. Our results indicate that MCC directly effects bladder cancer cells by inhibiting cellular proliferation through the induction of apoptosis, and has the potential for an indirect anti-cancer activity by stimulating cancer-infiltrating monocytes/macrophages to synthesize IL-12. © 1999 Cancer Research Campaig

Effect of silica nanoparticles on compressive properties of an epoxy polymer

The effect of nanosilica on compressive properties of an Epikote 828 epoxy at room temperature was studied. A 40 wt% nanosilica/epoxy masterbatch (nanopox F400) was used to prepare a series of epoxy based nanocomposites with 5-25 wt% nanosilica content. Static uniaxial compression tests were conducted on cubic and cylindrical specimens to study the compressive stress-strain response, failure mechanisms and damage characteristics of the pure and nanomodified epoxy. It was found that the compressive stiffness and strength were improved with increasing nanosilica content without significant reduction in failure strain. The presence of nanosilica improved ductility and promoted higher plastic hardening behaviour after yielding in comparison with the unmodified resin system. This result suggested that nanoparticles introduced additional mechanisms of energy absorption to enhance the compressive properties without reducing the deformation to failure