Biomedical Engineering Reference
In-Depth Information
expression in animals. In particular, the use of bioluminescence imaging
(BLI) to monitor disease processes in living animals, allows visualizing
cellular events, such as cell migration, signal transduction, proliferation
and apoptosis in the animal context (see (Hutchens and Luker 2007) for a
review). These technologies have been developed by using sensitive charge
coupled device (CCD) cameras to detect low levels of light emitted from
luciferase reporters
in vivo
. Although fl uorescence is almost universally
preferred over luminescence for image analysis of microscopic structure,
the latter presents some advantages. The main limitation of fl uorescence is
the higher backgrounds (due to a high infl ux of photons into the sample
that must be discriminated from the smaller emission of photons from
fl uorophores), leading to lower relative signals. The low background
inherent in luminescence is an advantage over fl uorescence since photons
are not required to create excited states.
In 1995, Contag and colleagues showed that it was possible to monitor
infectious processes in living animals using bioluminescence technology.
Initial studies were done with the bacterial pathogen
Salmonella enterica
transformed with the bacterial
lux
operon (Contag et al. 1995). Apart from
the well-known
in vivo
studies for visualization of cells targeted with
fl uorescent proteins (Barelle et al. 2004, Barelle et al. 2008, Del Poeta et al.
1999, Enjalbert et al. 2007, Gerami-Nejad et al. 2001), subsequent studies
have been developed by using
Photinus
and
Renilla
luciferases. The detection
of light
in vivo
is possible following the exogenous administration of the
substrates into the animal, luciferin and coelenterazine, respectively. In this
way, the fungal cells can be imaged inside the host animal and monitored
along the time by using CCD cameras. Recent studies have shown that the
signals of light emitted by the luciferases RLuc and FLuc can be detected
in animals. In the case of the fi refl y luciferase, Doyle and colleagues have
shown that the transcriptional fusion
ENO1
promoter-FLuc integrated in
C. albicans
can be detected in animals with vulvo-vaginal candidiasis (Doyle
et al. 2006b, Doyle et al. 2006a).
Bioluminescent
A. fumigatus
represents another model to study fungal
infections
in vivo
. In this case the promoter of glyceraldehide-3-phosphate
dehydrogenase is fused to FLuc in order study
in vitro
effectiveness of
drugs, the disease development, localization and burden of the fungal
pathogen within tissues, providing a suitable tool to study the effectiveness
of antifungals
in vivo
(Brock et al. 2008). However, these
in vivo
luminescent
reporters present some limitations. One of them is the limited permeability
of cells to the substrate luciferin and, the other one, the low signals detected
in deep organs by overlaying tissues (Doyle et al. 2006a). In order to avoid
these limitations, a recent study has described a new tool that implicates
an improved genetic construction by using a codon-optimized version of
