VEGF-C gene therapy augments postnatal lymphangiogenesis and ameliorates secondary lymphedema
Young-sup Yoon, Toshinori Murayama, Edwin Gravereaux, Tengiz Tkebuchava, Marcy Silver, Cynthia Curry, Andrea Wecker, Rudolf Kirchmair, Chun Song Hu, Marianne Kearney, Alan Ashare, David G. Jackson, Hajime Kubo, Jeffrey M. Isner, Douglas W. Losordo
Young-sup Yoon, Toshinori Murayama, Edwin Gravereaux, Tengiz Tkebuchava, Marcy Silver, Cynthia Curry, Andrea Wecker, Rudolf Kirchmair, Chun Song Hu, Marianne Kearney, Alan Ashare, David G. Jackson, Hajime Kubo, Jeffrey M. Isner, Douglas W. Losordo
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Article Genetics

VEGF-C gene therapy augments postnatal lymphangiogenesis and ameliorates secondary lymphedema

Abstract

Although lymphedema is a common clinical condition, treatment for this disabling condition remains limited and largely ineffective. Recently, it has been reported that overexpression of VEGF-C correlates with increased lymphatic vessel growth (lymphangiogenesis). However, the effect of VEGF-C–induced lymphangiogenesis on lymphedema has yet to be demonstrated. Here we investigated the impact of local transfer of naked plasmid DNA encoding human VEGF-C (phVEGF-C) on two animal models of lymphedema: one in the rabbit ear and the other in the mouse tail. In a rabbit model, following local phVEGF-C gene transfer, VEGFR-3 expression was significantly increased. This gene transfer led to a decrease in thickness and volume of lymphedema, improvement of lymphatic function demonstrated by serial lymphoscintigraphy, and finally, attenuation of the fibrofatty changes of the skin, the final consequences of lymphedema. The favorable effect of phVEGF-C on lymphedema was reconfirmed in a mouse tail model. Immunohistochemical analysis using lymphatic-specific markers: VEGFR-3, lymphatic endothelial hyaluronan receptor-1, together with the proliferation marker Ki-67 Ab revealed that phVEGF-C transfection potently induced new lymphatic vessel growth. This study, we believe for the first time, documents that gene transfer of phVEGF-C resolves lymphedema through direct augmentation of lymphangiogenesis. This novel therapeutic strategy may merit clinical investigation in patients with lymphedema.

Authors

Young-sup Yoon, Toshinori Murayama, Edwin Gravereaux, Tengiz Tkebuchava, Marcy Silver, Cynthia Curry, Andrea Wecker, Rudolf Kirchmair, Chun Song Hu, Marianne Kearney, Alan Ashare, David G. Jackson, Hajime Kubo, Jeffrey M. Isner, Douglas W. Losordo

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Temporal changes of lymphatic function visualized by lymphoscintigraphy....
Temporal changes of lymphatic function visualized by lymphoscintigraphy. (a and b) Orientation of the lymphoscintigraphic images. In normal ears, lymphatic flow assumes a linear pattern and the draining LNs are clearly visible. In the operated ear, the lymphatic passages were blocked, resulting in backward diffusion and no visualization of LNs. (c and d) Temporal changes in the saline group. Even at 12 weeks (d), lymphoscintigraphy demonstrates substantial impairment of lymphatic drainage of the saline-injected ear, indicated by dermal backflow and faint visualization of the LNs. (e and f) Temporal changes in the VEGF-C group. In the phVEGF-C–transfected ears, there was remarkable improvement of draining function. At 12 weeks, a linear passage of radiotracer, decreased dermal backflow, and increased uptake by LNs were observed. (g and h) Representative lymphoscintigraphic images and calculation of radioactivity index from the saline (g) and VEGF-C group (h). To quantitatively compare lymphatic drainage, the radioactivity within the ear was counted. Net radioactivity of the ear was obtained by subtracting γ counts at injection sites (arrows) from the total counts of the ear. The radioactivity index is the ratio of radioactivity of the operated ear divided by the radioactivity of the normal ear; this was used to compare lymphatic drainage function of the lymphedema ears. Higher ratios indicate more persistent radioactivity and less lymphatic drainage. (i) Comparison between the saline and VEGF-C groups shows the values were consistently lower in the VEGF-C group at 4, 8, and 12 weeks. *P < 0.05; **P < 0.01.