A double-labeled transgenic mouse was created
in our lab. The male ACTB-EGFP mice (Jackson Lab) mouse was cross-bred with a
homozygote mBSP9.0Luc female mouse . We named the mice with both Luc and GFP
genes as mBSP9.0Luc/β-ACT-EGFP. The production of the mouse line and the unique
features of the cells are illustrated in Figure
1A. Using the Xenogen 200 IVIS Imager we could identify pups that were
positive for both genes in the offsprings (Figure
1B, left panel). Bone marrow stem cells (BMSCs) isolated from
mBSP9.0Luc/β-ACT-EGFP was cultured and examined . BMSCs possess the
pluripotency to differentiate into a variety of cell types, including
osteoblasts, chondrocytes, and adipocytes. The cells were cultured in
osteogenic medium showing luciferase expression after introduction of luciferin
substrate indicating an osteogenic differentiation of the BMSCs. Non-osteogenic
treated cells only showed GFP fluorescent light (Figure 1B, right panel).
To test the sensitivity of the IVIS system in
detecting signals in a live animal we have performed another round of pilot
study using the BMSCs isolated from mBSP9.0Luc/β-ACT-EGFP transgenic mice.
After transplantation into the calvarial and mandibular defects in nude mice
for 2 and 7 days, respectively, we anesthetized the mice and placed them in the
Xenogen imager and luciferase and EGFP
expressions were measured using an IVIS Imaging System 200 Series in the live
animals as we did
previously [3,4]. Shown in Figure 2
were the representative images displaying strong luciferase and GFP signals in
the wound sites. These double-labeled
cells are unique and useful for bone tissue engineering and regeneration
studies. One genetic marker is a luciferase reporter gene driven by a mouse BSP
promoter that regulates the expression of BSP that is specific to mineralized
tissues [1,5-7]. The other genetic marker is GFP driven by a β-actin promoter
and a cytomegalovirus enhancer. By scanning the mouse using Xenogen IVIS in vivo imaging system which allows
real-time, non-invasive exploration of genes and cells in living animals, GFP
imaging is used to track the fate and migration of transplanted BMSCs, whereas
luciferase imaging serves as a marker for osteogenic differentiation of the
transplanted BMSCs because the luciferase-mediated bioluminescence will only be
appearing in newly formed bones. This
mouse model will greatly facilitate stem cells research as it permits the
determination of where these cells originate and how they undergo osteogenic
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