Citation Details

Journal Article

Adenoviridae/genetics, Anesthesia, Animals, Firefly Luciferin/administration & dosage/pharmacology, *Gene Expression Regulation/drug effects, Genetic Vectors/genetics, Luciferases/metabolism, Luminescent Measurements/*methods, Mice, Photons, Whole Body Imaging/*methods IVIS, Xenogen, Xen5

Molecular imaging offers many unique opportunities to study biological processes in intact organisms. Bioluminescence is the emission of light from biochemical reactions that occur within a living organism. Luciferase has been used as a reporter gene in transgenic mice but, until bioluminescence imaging was described, the detection of luciferase activity required either sectioning of the animal or excision of tissue and homogenization to measure enzyme activities in a conventional luminometer. Bioluminescence imaging (BLI) is based on the idea that biological light sources can be incorporated into cells and animal models artificially that does not naturally express the luminescent genes. This imaging modality has proven to be a very powerful methodology to detect luciferase reporter activity in intact animal models. This form of optical imaging is low cost and noninvasive and facilitates real-time analysis of disease processes at the molecular level in living organisms. Bioluminescence provides a noninvasive method to monitor gene expression in vivo and has enormous potential to elucidate the pathobiology of lung diseases in intact mouse models, including models of inflammation/injury, infection, and cancer.

Journal Article

Animals, Diagnostic Imaging/ methods, Female, Mice, Microscopy, Electron, Scanning, Photons, Proteus Infections/ diagnosis, Proteus mirabilis/drug effects/isolation & purification, Pseudomonas Infections/ diagnosis, Pseudomonas aeruginosa/drug effects/isolation & purification, Urinary Catheterization/ adverse effects, Urinary Tract Infections/ diagnosis IVIS, Xenogen, Xen5, Xen44

Urinary tract infections (UTIs) are among the most common bacterial infections acquired by humans, particularly in catheterized patients. A major problem with catheterization is the formation of bacterial biofilms on catheter material and the risk of developing persistent UTIs that are difficult to monitor and eradicate. To better understand the course of UTIs and allow more accurate studies of in vivo antibiotic efficacy, we developed a catheter-based biofilm infection model with mice, using bioluminescently engineered bacteria. Two important urinary tract pathogens, Pseudomonas aeruginosa and Proteus mirabilis, were made bioluminescent by stable insertion of a complete lux operon. Segments of catheter material (precolonized or postimplant infected) with either pathogen were placed transurethrally in the lumen of the bladder by using a metal stylet without surgical manipulation. The bioluminescent strains were sufficiently bright to be readily monitored from the outside of infected animals, using a low-light optical imaging system, including the ability to trace the ascending pattern of light-emitting bacteria through ureters to the kidneys. Placement of the catheter in the bladder not only resulted in the development of strong cystitis that persisted significantly longer than in mice challenged with bacterial suspensions alone but also required prolonged antibiotic treatment to reduce the level of infection. Treatment of infected mice for 4 days with ciprofloxacin at 30 mg/kg of body weight twice a day cured cystitis and renal infection in noncatheterized mice. Similarly, ciprofloxacin reduced the bacterial burden to undetectable levels in catheterized mice but did not inhibit rebound of the infection upon cessation of antibiotic therapy. This methodology easily allows spatial information to be monitored sequentially throughout the entire disease process, including ascending UTI, treatment efficacy, and relapse, all without exogenous sampling, which is not possible with conventional methods.

Journal Article

IVIS, Xenogen, Xen10

N/A

Journal Article

Animals, Cell Line, Cell Line, Transformed, Humans, Macrophages, Alveolar/*drug effects/immunology/*microbiology/pathology, Mice, Mice, Inbred C57BL, Morphine/administration & dosage/*therapeutic use, NF-kappa B/*antagonists & inhibitors/physiology, Neutrophil Infiltration/drug effects/immunology, Pneumonia, Pneumococcal/*drug therapy/*immunology/microbiology/mortality, Signal Transduction/*drug effects/immunology, Streptococcus pneumoniae/drug effects/*immunology, Time Factors, Toll-Like Receptor 2/physiology, Toll-Like Receptor 4/physiology, Toll-Like Receptor 9/*antagonists & inhibitors/physiology IVIS, Xenogen, Xen10

Resident alveolar macrophages and respiratory epithelium constitutes the first line of defense against invading lung pneumococci. Results from our study showed that increased mortality and bacterial outgrowth and dissemination seen in morphine-treated mice were further exaggerated following depletion of alveolar macrophages with liposomal clodronate. Using an in vitro alveolar macrophages and lung epithelial cells infection model, we show significant release of MIP-2 from alveolar macrophages, but not from lung epithelial cells, following 4 h of exposure of cells to pneumococci infection. Morphine treatment reduced MIP-2 release in pneumococci stimulated alveolar macrophages. Furthermore, morphine treatment inhibited Streptococcus pneumoniae-induced NF-kappaB-dependent gene transcription in alveolar macrophages following 2 h of in vitro infection. S. pneumoniae infection resulted in a significant induction of NF-kappaB activity only in TLR9 stably transfected HEK 293 cells, but not in TLR2 and TLR4 transfected HEK 293 cells, and morphine treatment inhibited S. pneumoniae-induced NF-kappaB activity in these cells. Moreover, morphine treatment also decreased bacterial uptake and killing in alveolar macrophages. Taken together, these results suggest that morphine treatment impairs TLR9-NF-kappaB signaling and diminishes bacterial clearance following S. pneumoniae infection in resident macrophages during the early stages of infection, leading to a compromised innate immune response.

Journal Article

Animals, Diagnostic Imaging, Disease Models, Animal, Female, Luminescent Measurements/methods, Meningitis, Pneumococcal/drug therapy/microbiology/ radiography, Mice, Mice, Inbred BALB C, Streptococcus pneumoniae/drug effects IVIS, Xenogen, Xen10

Noninvasive real-time in vivo bioluminescent imaging was used to assess the spread of Streptococcus pneumoniae throughout the spinal cord and brain during the acute stages of bacterial meningitis. A mouse model was established by lumbar (LP) or intracisternal (IC) injection of bioluminescent S. pneumoniae into the subarachnoid space. Bacteria replicated initially at the site of inoculation and spread progressively from the spinal cord to the brain or from the brain down to the cervical part of the spinal column and to the lower vertebral levels. After 24 hr, animals showed strong bioluminescent signals throughout the spinal canal, indicating acute meningitis of the intracranial and intraspinal meninges. A decline in bacterial cell viability, as judged by a reduction in the bioluminescent signal, was observed over time in animals treated with ceftriaxone, but not in untreated groups. Mice treated with the antibiotic survived infection, whereas all mice in untreated groups became moribund, first in the IC group then in the LP group. No untreated animal survived beyond 48 hr after induction of infection. Colony counts of infected cerebrospinal fluid (CSF) correlated positively with bioluminescent signals. This methodology is especially appealing because it allows detecting infected mice as early as 3 hr after inoculation, provide temporal, sequential, and spatial distribution of bacteria within the brain and spinal cord throughout the entire disease process and the rapid monitoring of treatment efficacy in a nondestructive manner. Moreover, it avoids the need to sacrifice the animals for CSF sampling and the potential manipulative damage that can occur with other conventional methods.