Fluorescent Proteins as Visible In Vivo Sensors - Progress in Molecular Biology and Translational Science - page 390

as in vitro. 18 To obtain the dual-color cells, RFP was expressed in the cy-
toplasm of cancer cells, and GFP linked to histone H2B was expressed in
the nucleus. Nuclear GFP expression enabled visualization of nuclear dy-
namics, whereas simultaneous cytoplasmic RFP expression enabled visu-
alization of nuclear-cytoplasmic ratios as well as simultaneous cell and
nuclear shape changes. The cell-cycle position of individual living cells
was readily visualized by the nuclear-cytoplasmic ratio and nuclear mor-
phology. Real-time induction of apoptosis was observed by nuclear size
changes and progressive nuclear fragmentation. Mitotic cells were visual-
ized by whole-body imaging after injection into the mouse ear. Common
carotid artery injection of dual-color cells and a reversible skin flap enabled
the external visualization of the dual-color cells in microvessels in the
mousebrainwhereextremeelongationofthecellbodyaswellasthenu-
cleus occurred.
2.2. Noninvasive cellular and subcellular imaging of cancer
cell - stromal cell interaction
To noninvasively image cancer cell/stromal cell interaction in the tumor
microenvironment (TME) anddrug response at the cellular level in live animals
in real time, we developed a new imageable three-color animal model. 19 The
model consists of GFP-expressing mice transplanted with dual-color cancer
cells labeled with GFP in the nucleus and RFP in the cytoplasm. An Olympus
IV100 laser scanning microscope, with ultra-narrow microscope objectives
(stick objectives), was used for three-color whole-body imaging of the two-
color cancer cells interacting with the GFP-expressing stromal cells. Drug re-
sponse of both cancer and stromal cells in the intact live animal was also imaged
in real time. Various in vivo phenomena of tumor-host interaction and cellular
dynamics were noninvasively imaged, including mitotic and apoptotic tumor
cells, stromal cells interacting with cancer cells, tumor vasculature, and tumor
blood flow ( Fig. 10.2 ).
2.3. Noninvasive cellular imaging in graft versus host disease
Noninvasive in vivo fluorescence imaging of the ear pinna enabled visualiza-
tion of GFP donor cells at the single-cell level in real time after allogeneic
hematopoietic stem-cell transplantation (HSCT). 20 Movement of donor
cells was increased by treatment with croton oil as an inflammatory reagent.
Treatment with dexamethasone, as an anti-inflammatory reagent, suppressed
donor cell infiltration.