Citation Details

Journal Article

Xen41, Xen 41, Pseudomonas aeruginosa Xen41, IVIS

BACKGROUND: : Experimental models of intestinal ischemia-reperfusion (IIR) injury are invariably performed in mice harboring their normal commensal flora, even though multiple IIR events occur in humans during prolonged intensive care confinement when they are colonized by a highly pathogenic hospital flora. The aims of this study were to determine whether the presence of the human pathogen Pseudomonas aeruginosa in the distal intestine potentiates the lethality of mice exposed to IIR and to determine what role any in vivo virulence activation plays in the observed mortality. METHODS: : Seven- to 9-week-old C57/BL6 mice were exposed to 15 minutes of superior mesenteric artery occlusion (SMAO) followed by direct intestinal inoculation of 1.0 x 10 colony-forming unit of P. aeruginosa PAO1 into the ileum and observed for mortality. Reiterative studies were performed in separate groups of mice to evaluate both the migration/dissemination pattern and in vivo virulence activation of intestinally inoculated strains using live photon camera imaging of both a constitutive bioluminescent P. aeruginosa PAO1 derivative XEN41 and an inducible reporter derivative of PAO1, the PAO1/lecA:luxCDABE that conditionally expresses the quorum sensing-dependent epithelial disrupting virulence protein PA 1 Lectin (PA-IL). RESULTS: : Mice exposed to 15 minutes of SMAO and reperfusion with intestinal inoculation of P. aeruginosa had a significantly increased mortality rate (p < 0.001) of 100% compared with <10% for sham-operated mice intestinally inoculated with P. aeruginosa without SMAO and IIR alone (<50%). Migration/dissemination patterns of P. aeruginosa in mice subjected to IIR demonstrated proximal migration of distally injected strains and translocation to mesenteric lymph nodes, liver, spleen, lung, and kidney. A key role for in vivo virulence expression of the barrier disrupting adhesin PA-IL during IIR was established since its expression was enhanced during IR and mutant strains lacking PA-IL displayed attenuated mortality. CONCLUSIONS: : The presence of intestinal P. aeruginosa potentiates the lethal effect of IIR in mice in part due to in vivo virulence activation of its epithelial barrier disrupting protein PA-IL.

Journal Article

Xen36, Xen 36, Staphylococcus aureus Xen36, IVIS, Animals; Arthroplasty; Biofilms/growth & development; Bone Wires/microbiology; Interleukin-1beta/*metabolism; Male; Mice; Mice, Congenic; Mice, Inbred C57BL; Myeloid Differentiation Factor 88/metabolism; Neutrophil Infiltration; Prosthesis-Related Infections/*immunology/metabolism; Staphylococcal Infections/*immunology/metabolism; Staphylococcus aureus; Toll-Like Receptor 2/*metabolism

MyD88 is an adapter molecule that is used by both IL-1R and TLR family members to initiate downstream signaling and promote immune responses. Given that IL-1beta is induced after Staphylococcus aureus infections and TLR2 is activated by S. aureus lipopeptides, we hypothesized that IL-1beta and TLR2 contribute to MyD88-dependent protective immune responses against post-arthroplasty S. aureus infections. To test this hypothesis, we used a mouse model of a post-arthroplasty S. aureus infection to compare the bacterial burden, biofilm formation and neutrophil recruitment in IL-1beta-deficient, TLR2-deficient and wild-type (wt) mice. By using in vivo bioluminescence imaging, we found that the bacterial burden in IL-1beta-deficient mice was 26-fold higher at 1 day after infection and remained 3- to 10-fold greater than wt mice through day 42. In contrast, the bacterial burden in TLR2-deficient mice did not differ from wt mice. In addition, implants harvested from IL-1beta-deficient mice had more biofilm formation and 14-fold higher adherent bacteria compared with those from wt mice. Finally, IL-1beta-deficient mice had approximately 50% decreased neutrophil recruitment to the infected postoperative joints than wt mice. Taken together, these findings suggest a mechanism by which IL-1beta induces neutrophil recruitment to help control the bacterial burden and the ensuing biofilm formation in a post-surgical joint.

Journal Article

Animals; Anthrax; Bacillus anthracis; Bioware; Disease Models, Animal; Gastrointestinal Diseases; Inhalation Exposure; Luciferases; Luminescence; Luminescent Measurements; Lymph Nodes; Mice; Mice, Inbred BALB C; Nasal Cavity; Organisms, Genetically Modified; Peyer's Patches; Pharynx; pXen-5; Skin; Spores, Bacterial

Bacillus anthracis causes three forms of anthrax: inhalational, gastrointestinal, and cutaneous. Anthrax is characterized by both toxemia, which is caused by secretion of immunomodulating toxins (lethal toxin and edema toxin), and septicemia, which is associated with bacterial encapsulation. Here we report that, contrary to the current view of B. anthracis pathogenesis, B. anthracis spores germinate and establish infections at the initial site of inoculation in both inhalational and cutaneous infections without needing to be transported to draining lymph nodes, and that inhaled spores establish initial infection in nasal-associated lymphoid tissues. Furthermore, we found that Peyer's patches in the mouse intestine are the primary site of bacterial growth after intragastric inoculation, thus establishing an animal model of gastrointestinal anthrax. All routes of infection progressed to the draining lymph nodes, spleen, lungs, and ultimately the blood. These discoveries were made possible through the development of a novel dynamic mouse model of B. anthracis infection using bioluminescent non-toxinogenic capsulated bacteria that can be visualized within the mouse in real-time, and demonstrate the value of in vivo imaging in the analysis of B. anthracis infection. Our data imply that previously unrecognized portals of bacterial entry demand more intensive investigation, and will significantly transform the current perception of inhalational, gastrointestinal, and cutaneous B. anthracis pathogenesis.

Journal Article

IVIS, B16-F10-luc-G5, B16F10-luc-G5, B16-F10-luc, B16F10-luc,

Tumor progression is associated with the release of signaling substances from the primary tumor into the bloodstream. Tumor-derived cytokines are known to promote the mobilization and the recruitment of cells from the bone marrow, including endothelial progenitor cells (EPC). Here, we examined whether such paracrine influence could also influence the capacity of EPC to interfere with circulating metastatic cells. We therefore consecutively injected EPC pre-stimulated by tumor conditioned medium (CM-EPC) and luciferase-expressing B16 melanoma cells to mice. A net decrease in metastases spreading (vs non-stimulated EPC) led us to carry out a 2D-DIGE proteomic study to identify possible mediators of EPC-driven protection. Among 33 proteins exhibiting significant changes in expression, SPARC presented the highest induction after EPC exposure to CM. We then showed that contrary to control EPC, SPARC-silenced EPC were not able to reduce the extent of metastases when injected with B16 melanoma cells. Using adhesion tests and the hanging drop assay, we further documented that cell-cell interactions between CM-EPC and melanoma cells were promoted in a SPARC-dependent manner. This interaction led to the engulfment of melanoma cells by CM-EPC, a process prevented by SPARC silencing and mimicked by recombinant SPARC. Finally, we showed that contrary to melanoma cells, the pro-metastatic human breast cancer cell line MDA-MB231-D3H2 reduced SPARC expression in human EPC and stimulated metastases spreading. Our findings unravel the influence of tumor cells on EPC phenotypes through a SPARC-driven accentuation of macrophagic capacity associated with limitations to metastatic spread.

Journal Article

IntegriSense, Animals; Bone Neoplasms/*diagnosis/*secondary; Diagnostic Imaging/*methods; Disease Models, Animal; Disease Progression; Humans; Molecular Imaging/methods; Neoplasm Metastasis/diagnosis

Bone metastasis is a complex process that ultimately leads to devastating metastatic bone disease. It is therefore of key interest to unravel the mechanisms underlying the multistep process of skeletal metastasis and cancer-induced bone disease, and to develop better treatment and management of patients with this devastating disease. Fortunately, novel technologies are rapidly emerging that allow real-time imaging of molecules, pathogenic processes, drug delivery and drug response in preclinical in vivo models. The outcome of these experimental studies will facilitate clinical cancer research by improving the detection of cancer cell invasion, metastasis and therapy response.