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Imaging In Mice With Fluorescent Proteins: From Macro To Subcellular

View Article: PubMed Central - PubMed

ABSTRACT

Whole-body imaging with fluorescent proteins has been shown to be a powerful technology with many applications in small animals. Brighter, red-shifted proteins can make whole-body imaging even more sensitive due to reduced absorption by tissues and less scatter. For example, a new protein called Katushka has been isolated that is the brightest known protein with emission at wavelengths longer than 620 nm. This new protein offers potential for noninvasive whole-body macro imaging such as of tumor growth. For subcellular imaging, to observe cytoplasmic and nuclear dynamics in the living mouse, cancer cells were labeled in the nucleus with green fluorescent protein and with red fluorescent protein in the cytoplasm. The nuclear and cytoplasmic behavior of cancer cells in real time in blood vessels was imaged as they trafficked by various means or adhered to the vessel surface in the abdominal skin flap. During extravasation, real-time dual-color imaging showed that cytoplasmic processes of the cancer cells exited the vessels first, with nuclei following along the cytoplasmic projections. Both cytoplasm and nuclei underwent deformation during extravasation. Cancer cells trafficking in lymphatic vessels was also imaged. To noninvasively image cancer cell/stromal cell interaction in the tumor microenvironment as well as drug response at the cellular level in live animals in real time, we developed a new imageable three-color animal model. The model consists of GFP-expressing mice transplanted with the dual-color cancer cells. With the dual-color cancer cells and a highly sensitive small animal imaging system, subcellular dynamics can now be observed in live mice in real time. Fluorescent proteins thus enable both macro and micro imaging technology and thereby provide the basis for the new field of in vivo cell biology.

No MeSH data available.


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Stable high GFP- and RFP-expressing dual-color cancer cells in vitroMouse mammary tumor (MMT) cells were initially transduced with RFP and the neomycin resistance gene. The cells were subsequently transduced with histone H2B-GFP and the hygromycin resistance gene. Double transformants were selected with G418 and hygromycin, and stable clones were established. Bar = 50 μm [31].
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f3-sensors-08-01157: Stable high GFP- and RFP-expressing dual-color cancer cells in vitroMouse mammary tumor (MMT) cells were initially transduced with RFP and the neomycin resistance gene. The cells were subsequently transduced with histone H2B-GFP and the hygromycin resistance gene. Double transformants were selected with G418 and hygromycin, and stable clones were established. Bar = 50 μm [31].

Mentions: To noninvasively visualize cellular and subcellular events in the tumor microenvironment in real time in the live mouse, we used the Olympus IV100 laser scanning microscope with a 0.3-mm-diameter stick objective that is up to 2 cm in length. The system allows an imaging depth that extends to at least 200 μm. This novel imaging system, coupled with the use of the cancer cells labeled in the nucleus with GFP and RFP in the cytoplasm (Figure 3) and a transgenic GFP or RFP mouse as the host (Figure 5), has enabled noninvasive in vivo imaging of the cancer and stromal cells in the tumor microenvironment at the subcellular level. Three-color whole-body imaging of the two-color cancer cells interacting with the GFP-expressing stromal cells was carried out. In this model, drug response 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 imaged, including mitotic and apoptotic cancer cells, stromal cells intimately interacting with the cancer cells, tumor vasculature, and tumor blood flow (Fig. 4). This new model system enabled the first cellular and subcellular images of unperturbed tumors in the live intact animal. New visible real-time targets for novel anticancer agents are provided in this model, including the color-coded interacting cancer and stromal cells, tumor vasculature, and blood flow. This imageable model should lead to many new insights of the tumor microenvironment [10].


Imaging In Mice With Fluorescent Proteins: From Macro To Subcellular
Stable high GFP- and RFP-expressing dual-color cancer cells in vitroMouse mammary tumor (MMT) cells were initially transduced with RFP and the neomycin resistance gene. The cells were subsequently transduced with histone H2B-GFP and the hygromycin resistance gene. Double transformants were selected with G418 and hygromycin, and stable clones were established. Bar = 50 μm [31].
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3927509&req=5

f3-sensors-08-01157: Stable high GFP- and RFP-expressing dual-color cancer cells in vitroMouse mammary tumor (MMT) cells were initially transduced with RFP and the neomycin resistance gene. The cells were subsequently transduced with histone H2B-GFP and the hygromycin resistance gene. Double transformants were selected with G418 and hygromycin, and stable clones were established. Bar = 50 μm [31].
Mentions: To noninvasively visualize cellular and subcellular events in the tumor microenvironment in real time in the live mouse, we used the Olympus IV100 laser scanning microscope with a 0.3-mm-diameter stick objective that is up to 2 cm in length. The system allows an imaging depth that extends to at least 200 μm. This novel imaging system, coupled with the use of the cancer cells labeled in the nucleus with GFP and RFP in the cytoplasm (Figure 3) and a transgenic GFP or RFP mouse as the host (Figure 5), has enabled noninvasive in vivo imaging of the cancer and stromal cells in the tumor microenvironment at the subcellular level. Three-color whole-body imaging of the two-color cancer cells interacting with the GFP-expressing stromal cells was carried out. In this model, drug response 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 imaged, including mitotic and apoptotic cancer cells, stromal cells intimately interacting with the cancer cells, tumor vasculature, and tumor blood flow (Fig. 4). This new model system enabled the first cellular and subcellular images of unperturbed tumors in the live intact animal. New visible real-time targets for novel anticancer agents are provided in this model, including the color-coded interacting cancer and stromal cells, tumor vasculature, and blood flow. This imageable model should lead to many new insights of the tumor microenvironment [10].

View Article: PubMed Central - PubMed

ABSTRACT

Whole-body imaging with fluorescent proteins has been shown to be a powerful technology with many applications in small animals. Brighter, red-shifted proteins can make whole-body imaging even more sensitive due to reduced absorption by tissues and less scatter. For example, a new protein called Katushka has been isolated that is the brightest known protein with emission at wavelengths longer than 620 nm. This new protein offers potential for noninvasive whole-body macro imaging such as of tumor growth. For subcellular imaging, to observe cytoplasmic and nuclear dynamics in the living mouse, cancer cells were labeled in the nucleus with green fluorescent protein and with red fluorescent protein in the cytoplasm. The nuclear and cytoplasmic behavior of cancer cells in real time in blood vessels was imaged as they trafficked by various means or adhered to the vessel surface in the abdominal skin flap. During extravasation, real-time dual-color imaging showed that cytoplasmic processes of the cancer cells exited the vessels first, with nuclei following along the cytoplasmic projections. Both cytoplasm and nuclei underwent deformation during extravasation. Cancer cells trafficking in lymphatic vessels was also imaged. To noninvasively image cancer cell/stromal cell interaction in the tumor microenvironment as well as drug response at the cellular level in live animals in real time, we developed a new imageable three-color animal model. The model consists of GFP-expressing mice transplanted with the dual-color cancer cells. With the dual-color cancer cells and a highly sensitive small animal imaging system, subcellular dynamics can now be observed in live mice in real time. Fluorescent proteins thus enable both macro and micro imaging technology and thereby provide the basis for the new field of in vivo cell biology.

No MeSH data available.


Related in: MedlinePlus