Limits...
Real-time monitoring of tumorigenesis, dissemination, & drug response in a preclinical model of lymphangioleiomyomatosis/tuberous sclerosis complex.

Liu F, Lunsford EP, Tong J, Ashitate Y, Gibbs SL, Yu J, Choi HS, Henske EP, Frangioni JV - PLoS ONE (2012)

Bottom Line: Surprisingly, TSC2-deficient cells administered intratracheally resulted in rapid dissemination to lymph node basins throughout the body, and histopathological changes in the lung consistent with LAM.Estrogen was found to be permissive for tumor growth and dissemination.We generated homogeneous NIS/GFP co-expressing TSC2-deficient, patient-derived cells that can proliferate and migrate in vivo after intratracheal instillation.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT

Background: TSC2-deficient cells can proliferate in the lungs, kidneys, and other organs causing devastating progressive multisystem disorders such as lymphangioleiomyomatosis (LAM) and tuberous sclerosis complex (TSC). Preclinical models utilizing LAM patient-derived cells have been difficult to establish. We developed a novel animal model system to study the molecular mechanisms of TSC/LAM pathogenesis and tumorigenesis and provide a platform for drug testing.

Methods and findings: TSC2-deficient human cells, derived from the angiomyolipoma of a LAM patient, were engineered to co-express both sodium-iodide symporter (NIS) and green fluorescent protein (GFP). Cells were inoculated intraparenchymally, intravenously, or intratracheally into athymic NCr nu/nu mice and cells were tracked and quantified using single photon emission computed tomography (SPECT) and computed tomography (CT). Surprisingly, TSC2-deficient cells administered intratracheally resulted in rapid dissemination to lymph node basins throughout the body, and histopathological changes in the lung consistent with LAM. Estrogen was found to be permissive for tumor growth and dissemination. Rapamycin inhibited tumor growth, but tumors regrew after the drug treatment was withdrawn.

Conclusions: We generated homogeneous NIS/GFP co-expressing TSC2-deficient, patient-derived cells that can proliferate and migrate in vivo after intratracheal instillation. Although the animal model we describe has some limitations, we demonstrate that systemic tumors formed from TSC2-deficient cells can be monitored and quantified noninvasively over time using SPECT/CT, thus providing a much needed model system for in vivo drug testing and mechanistic studies of TSC2-deficient cells and their related clinical syndromes.

Show MeSH

Related in: MedlinePlus

Proliferation and pathogenesis of TSC2-deficient cells: pulmonary LAM like-nodules and cysts.A. Ex vivo SPECT/CT imaging and antibody and H&E staining of lung. Resected whole lungs (left column) from LAM/TSC-bearing mice (top row) or control mice (bottom row) at 2 weeks postadministration of 621-327 cells. Scale bar = 1 mm. Also shown are H&E staining (middle column) and anti-GFP antibody staining (right column) of frozen sections from lungs at 4 weeks postadministration of 621-327 cells. Scale bars = 50 µm. B. H&E staining of paraffin-embedded lung tissue 15 weeks postadministration of 621-327 cells (top row) at low (top left) and high (top right and bottom row of dotted rectangles) magnification. Scale bars = 50 µm. C. Same as (B) except frozen sections at 26 weeks postadministration of 621-327 cells (left) with anti-human NIS antibody staining (right). Scale bars = 50 µm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3376142&req=5

pone-0038589-g004: Proliferation and pathogenesis of TSC2-deficient cells: pulmonary LAM like-nodules and cysts.A. Ex vivo SPECT/CT imaging and antibody and H&E staining of lung. Resected whole lungs (left column) from LAM/TSC-bearing mice (top row) or control mice (bottom row) at 2 weeks postadministration of 621-327 cells. Scale bar = 1 mm. Also shown are H&E staining (middle column) and anti-GFP antibody staining (right column) of frozen sections from lungs at 4 weeks postadministration of 621-327 cells. Scale bars = 50 µm. B. H&E staining of paraffin-embedded lung tissue 15 weeks postadministration of 621-327 cells (top row) at low (top left) and high (top right and bottom row of dotted rectangles) magnification. Scale bars = 50 µm. C. Same as (B) except frozen sections at 26 weeks postadministration of 621-327 cells (left) with anti-human NIS antibody staining (right). Scale bars = 50 µm.

Mentions: Resection of tissue with high radiotracer uptake (Figure S2) revealed a pattern consistent with lymphatic or hematogenous spread from the lung to lymph node basins. Indeed, histopathological analysis of lymph nodes at early time points after intratracheal administration revealed NIS−/GFP-expressing tumor deposits within lymph nodes (Figure 3A). Early tumor nodules were often seen within semi-encapsulated structures in lymph nodes (Figure 3B). By weeks 3 to 4, most lymph nodes analyzed were completely obliterated by the tumor cells with abundant or medium cytoplasm (arrow head). Interestingly, by 15 weeks postinoculation, some tumor cells developed a smooth muscle cell morphology, and some lymph nodes developed cystic or fluid-filled structures, although these cysts are not histologically identical to those observed in human LAM (Figure 3C). The weak SPECT signal measured in living mouse lungs (Figure 2A), and the strong signal seen after sacrifice (Figure 4A) suggest that the signal-to-noise ratio in the lungs was significantly degraded by respiratory motion.


Real-time monitoring of tumorigenesis, dissemination, & drug response in a preclinical model of lymphangioleiomyomatosis/tuberous sclerosis complex.

Liu F, Lunsford EP, Tong J, Ashitate Y, Gibbs SL, Yu J, Choi HS, Henske EP, Frangioni JV - PLoS ONE (2012)

Proliferation and pathogenesis of TSC2-deficient cells: pulmonary LAM like-nodules and cysts.A. Ex vivo SPECT/CT imaging and antibody and H&E staining of lung. Resected whole lungs (left column) from LAM/TSC-bearing mice (top row) or control mice (bottom row) at 2 weeks postadministration of 621-327 cells. Scale bar = 1 mm. Also shown are H&E staining (middle column) and anti-GFP antibody staining (right column) of frozen sections from lungs at 4 weeks postadministration of 621-327 cells. Scale bars = 50 µm. B. H&E staining of paraffin-embedded lung tissue 15 weeks postadministration of 621-327 cells (top row) at low (top left) and high (top right and bottom row of dotted rectangles) magnification. Scale bars = 50 µm. C. Same as (B) except frozen sections at 26 weeks postadministration of 621-327 cells (left) with anti-human NIS antibody staining (right). Scale bars = 50 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3376142&req=5

pone-0038589-g004: Proliferation and pathogenesis of TSC2-deficient cells: pulmonary LAM like-nodules and cysts.A. Ex vivo SPECT/CT imaging and antibody and H&E staining of lung. Resected whole lungs (left column) from LAM/TSC-bearing mice (top row) or control mice (bottom row) at 2 weeks postadministration of 621-327 cells. Scale bar = 1 mm. Also shown are H&E staining (middle column) and anti-GFP antibody staining (right column) of frozen sections from lungs at 4 weeks postadministration of 621-327 cells. Scale bars = 50 µm. B. H&E staining of paraffin-embedded lung tissue 15 weeks postadministration of 621-327 cells (top row) at low (top left) and high (top right and bottom row of dotted rectangles) magnification. Scale bars = 50 µm. C. Same as (B) except frozen sections at 26 weeks postadministration of 621-327 cells (left) with anti-human NIS antibody staining (right). Scale bars = 50 µm.
Mentions: Resection of tissue with high radiotracer uptake (Figure S2) revealed a pattern consistent with lymphatic or hematogenous spread from the lung to lymph node basins. Indeed, histopathological analysis of lymph nodes at early time points after intratracheal administration revealed NIS−/GFP-expressing tumor deposits within lymph nodes (Figure 3A). Early tumor nodules were often seen within semi-encapsulated structures in lymph nodes (Figure 3B). By weeks 3 to 4, most lymph nodes analyzed were completely obliterated by the tumor cells with abundant or medium cytoplasm (arrow head). Interestingly, by 15 weeks postinoculation, some tumor cells developed a smooth muscle cell morphology, and some lymph nodes developed cystic or fluid-filled structures, although these cysts are not histologically identical to those observed in human LAM (Figure 3C). The weak SPECT signal measured in living mouse lungs (Figure 2A), and the strong signal seen after sacrifice (Figure 4A) suggest that the signal-to-noise ratio in the lungs was significantly degraded by respiratory motion.

Bottom Line: Surprisingly, TSC2-deficient cells administered intratracheally resulted in rapid dissemination to lymph node basins throughout the body, and histopathological changes in the lung consistent with LAM.Estrogen was found to be permissive for tumor growth and dissemination.We generated homogeneous NIS/GFP co-expressing TSC2-deficient, patient-derived cells that can proliferate and migrate in vivo after intratracheal instillation.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT

Background: TSC2-deficient cells can proliferate in the lungs, kidneys, and other organs causing devastating progressive multisystem disorders such as lymphangioleiomyomatosis (LAM) and tuberous sclerosis complex (TSC). Preclinical models utilizing LAM patient-derived cells have been difficult to establish. We developed a novel animal model system to study the molecular mechanisms of TSC/LAM pathogenesis and tumorigenesis and provide a platform for drug testing.

Methods and findings: TSC2-deficient human cells, derived from the angiomyolipoma of a LAM patient, were engineered to co-express both sodium-iodide symporter (NIS) and green fluorescent protein (GFP). Cells were inoculated intraparenchymally, intravenously, or intratracheally into athymic NCr nu/nu mice and cells were tracked and quantified using single photon emission computed tomography (SPECT) and computed tomography (CT). Surprisingly, TSC2-deficient cells administered intratracheally resulted in rapid dissemination to lymph node basins throughout the body, and histopathological changes in the lung consistent with LAM. Estrogen was found to be permissive for tumor growth and dissemination. Rapamycin inhibited tumor growth, but tumors regrew after the drug treatment was withdrawn.

Conclusions: We generated homogeneous NIS/GFP co-expressing TSC2-deficient, patient-derived cells that can proliferate and migrate in vivo after intratracheal instillation. Although the animal model we describe has some limitations, we demonstrate that systemic tumors formed from TSC2-deficient cells can be monitored and quantified noninvasively over time using SPECT/CT, thus providing a much needed model system for in vivo drug testing and mechanistic studies of TSC2-deficient cells and their related clinical syndromes.

Show MeSH
Related in: MedlinePlus