Limits...
In vivo lymphatic imaging of a human inflammatory breast cancer model.

Agollah GD, Wu G, Sevick-Muraca EM, Kwon S - J Cancer (2014)

Bottom Line: Inflammatory breast cancer (IBC) remains the most aggressive type of breast cancer with the greatest potential for metastasis and as a result, the highest mortality rate.Herein, we non-invasively and longitudinally imaged lymphatics in an animal model of IBC using near-infrared fluorescence (NIRF) imaging.We also observed increased and dilated fluorescent lymphatic vessels in the tumor periphery, which was confirmed by ex vivo immunohistochemical staining of lymphatic vessels.

View Article: PubMed Central - PubMed

Affiliation: 1. Center for Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, The University of Texas Health Science Center, Houston, TX 77030; ; 2. The University of Texas Graduate School of Biomedical Sciences at Houston. The University of Texas MD Anderson Cancer Center, Houston, Texas 77030.

ABSTRACT

Background: Inflammatory breast cancer (IBC) remains the most aggressive type of breast cancer with the greatest potential for metastasis and as a result, the highest mortality rate. IBC cells invade and metastasize through dermal lymphatic vessels; however, it is unknown how lymphatic drainage patterns change during IBC growth and metastasis. Herein, we non-invasively and longitudinally imaged lymphatics in an animal model of IBC using near-infrared fluorescence (NIRF) imaging.

Materials and methods: Mice were imaged in vivo prior to, and up to 11 weeks after subcutaneous or orthotopic inoculation of human IBC SUM149 cells, which were stably transfected with infrared fluorescence protein (iRFP) gene reporter (SUM149-iRFP), following intradermal (i.d.) injection of indocyanine green (ICG).

Results: Fluorescence images showed well-defined lymphatic vessels prior to SUM149-iRFP inoculation. However, altered lymphatic drainage patterns including rerouting of lymphatic drainage were detected in mice with SUM149-iRFP, due to lymphatic obstruction of normal lymphatic drainages caused by tumor growth. In addition, we observed tortuous lymphatic vessels and extravasation of ICG-laden lymph in mice with SUM149-iRFP. We also observed increased and dilated fluorescent lymphatic vessels in the tumor periphery, which was confirmed by ex vivo immunohistochemical staining of lymphatic vessels.

Conclusions: Our pre-clinical studies demonstrate that non-invasive NIRF imaging can provide a method to assess changes in lymphatic drainage patterns during IBC growth and metastasis.

No MeSH data available.


Related in: MedlinePlus

White light, NIR, and iRFP fluorescent images prior to and for up to 9 weeks after s.c. inoculation of SUM149-iRFP cells. Images were acquired 10 mins after i.d. injection of ICG. NIR fluorescent images (green) of the lymphatics were fused with iRFP fluorescent images (red) of the gene reporters in the tumor. Magnified fluorescent images of the white rectangles were also acquired. Arrow, ILN. Double arrow, injection site. Arrow head, SUM149-iRFP tumor. Open arrow, lymphatic capillaries due to dermal backflow. Scale, 1 mm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4216802&req=5

Figure 4: White light, NIR, and iRFP fluorescent images prior to and for up to 9 weeks after s.c. inoculation of SUM149-iRFP cells. Images were acquired 10 mins after i.d. injection of ICG. NIR fluorescent images (green) of the lymphatics were fused with iRFP fluorescent images (red) of the gene reporters in the tumor. Magnified fluorescent images of the white rectangles were also acquired. Arrow, ILN. Double arrow, injection site. Arrow head, SUM149-iRFP tumor. Open arrow, lymphatic capillaries due to dermal backflow. Scale, 1 mm.

Mentions: To assess how a different implantation model would affect lymphatic drainage, we also performed subcutaneous (s.c) tumor implantation and longitudinally monitored tumor growth and changes in lymphatic drainage (Figure 4). NIRF imaging showed ICG-laden lymph draining along the lymphatic vessels in the skin above the tumor at 1 week p.i., with increased fluorescent intensity and fluorescent spots. At 3 weeks after inoculation, we observed ICG accumulation in the tumor as evidenced by strong fluorescence likely due to leaky lymphatic vessels (Additional file 4: video 4). However, ICG intratumoral accumulation was not detected at 4 weeks p.i.; instead, we observed dermal backflow proximal to the tumor. At 5 weeks p.i. alternate lymphatic routes were observed around the tumor which were dilated by 7 weeks p.i; and ICG accumulated anteriorly around the tumor margin at week 9. Furthermore, our in vivo imaging data also showed iRFP fluorescence in the TdILN in this mouse, indicative of LN metastasis (Figure 5A) and was confirmed by ex vivo intravital iRFP-fluorescence imaging of the TdILN. SUM149 overexpresses luminal cytokeratin 8 20; therefore, we further confirmed local LN metastasis by IHC staining and detected cytokeratin-positive cells in the TdILN compared to the contralateral ILN (Figure 5B).


In vivo lymphatic imaging of a human inflammatory breast cancer model.

Agollah GD, Wu G, Sevick-Muraca EM, Kwon S - J Cancer (2014)

White light, NIR, and iRFP fluorescent images prior to and for up to 9 weeks after s.c. inoculation of SUM149-iRFP cells. Images were acquired 10 mins after i.d. injection of ICG. NIR fluorescent images (green) of the lymphatics were fused with iRFP fluorescent images (red) of the gene reporters in the tumor. Magnified fluorescent images of the white rectangles were also acquired. Arrow, ILN. Double arrow, injection site. Arrow head, SUM149-iRFP tumor. Open arrow, lymphatic capillaries due to dermal backflow. Scale, 1 mm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4216802&req=5

Figure 4: White light, NIR, and iRFP fluorescent images prior to and for up to 9 weeks after s.c. inoculation of SUM149-iRFP cells. Images were acquired 10 mins after i.d. injection of ICG. NIR fluorescent images (green) of the lymphatics were fused with iRFP fluorescent images (red) of the gene reporters in the tumor. Magnified fluorescent images of the white rectangles were also acquired. Arrow, ILN. Double arrow, injection site. Arrow head, SUM149-iRFP tumor. Open arrow, lymphatic capillaries due to dermal backflow. Scale, 1 mm.
Mentions: To assess how a different implantation model would affect lymphatic drainage, we also performed subcutaneous (s.c) tumor implantation and longitudinally monitored tumor growth and changes in lymphatic drainage (Figure 4). NIRF imaging showed ICG-laden lymph draining along the lymphatic vessels in the skin above the tumor at 1 week p.i., with increased fluorescent intensity and fluorescent spots. At 3 weeks after inoculation, we observed ICG accumulation in the tumor as evidenced by strong fluorescence likely due to leaky lymphatic vessels (Additional file 4: video 4). However, ICG intratumoral accumulation was not detected at 4 weeks p.i.; instead, we observed dermal backflow proximal to the tumor. At 5 weeks p.i. alternate lymphatic routes were observed around the tumor which were dilated by 7 weeks p.i; and ICG accumulated anteriorly around the tumor margin at week 9. Furthermore, our in vivo imaging data also showed iRFP fluorescence in the TdILN in this mouse, indicative of LN metastasis (Figure 5A) and was confirmed by ex vivo intravital iRFP-fluorescence imaging of the TdILN. SUM149 overexpresses luminal cytokeratin 8 20; therefore, we further confirmed local LN metastasis by IHC staining and detected cytokeratin-positive cells in the TdILN compared to the contralateral ILN (Figure 5B).

Bottom Line: Inflammatory breast cancer (IBC) remains the most aggressive type of breast cancer with the greatest potential for metastasis and as a result, the highest mortality rate.Herein, we non-invasively and longitudinally imaged lymphatics in an animal model of IBC using near-infrared fluorescence (NIRF) imaging.We also observed increased and dilated fluorescent lymphatic vessels in the tumor periphery, which was confirmed by ex vivo immunohistochemical staining of lymphatic vessels.

View Article: PubMed Central - PubMed

Affiliation: 1. Center for Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, The University of Texas Health Science Center, Houston, TX 77030; ; 2. The University of Texas Graduate School of Biomedical Sciences at Houston. The University of Texas MD Anderson Cancer Center, Houston, Texas 77030.

ABSTRACT

Background: Inflammatory breast cancer (IBC) remains the most aggressive type of breast cancer with the greatest potential for metastasis and as a result, the highest mortality rate. IBC cells invade and metastasize through dermal lymphatic vessels; however, it is unknown how lymphatic drainage patterns change during IBC growth and metastasis. Herein, we non-invasively and longitudinally imaged lymphatics in an animal model of IBC using near-infrared fluorescence (NIRF) imaging.

Materials and methods: Mice were imaged in vivo prior to, and up to 11 weeks after subcutaneous or orthotopic inoculation of human IBC SUM149 cells, which were stably transfected with infrared fluorescence protein (iRFP) gene reporter (SUM149-iRFP), following intradermal (i.d.) injection of indocyanine green (ICG).

Results: Fluorescence images showed well-defined lymphatic vessels prior to SUM149-iRFP inoculation. However, altered lymphatic drainage patterns including rerouting of lymphatic drainage were detected in mice with SUM149-iRFP, due to lymphatic obstruction of normal lymphatic drainages caused by tumor growth. In addition, we observed tortuous lymphatic vessels and extravasation of ICG-laden lymph in mice with SUM149-iRFP. We also observed increased and dilated fluorescent lymphatic vessels in the tumor periphery, which was confirmed by ex vivo immunohistochemical staining of lymphatic vessels.

Conclusions: Our pre-clinical studies demonstrate that non-invasive NIRF imaging can provide a method to assess changes in lymphatic drainage patterns during IBC growth and metastasis.

No MeSH data available.


Related in: MedlinePlus