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Quantitative measurement of lymphatic function in mice by noninvasive near-infrared imaging of a peripheral vein
Steven T. Proulx, … , Jean-Christophe Leroux, Michael Detmar
Steven T. Proulx, … , Jean-Christophe Leroux, Michael Detmar
Published January 12, 2017
Citation Information: JCI Insight. 2017;2(1):e90861. https://doi.org/10.1172/jci.insight.90861.
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Resource and Technical Advance Vascular biology

Quantitative measurement of lymphatic function in mice by noninvasive near-infrared imaging of a peripheral vein

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Abstract

Optical imaging methods have been developed to measure lymphatic function in skin; however, the lymphatic system of many organs is not accessible to this technology. Since lymphatic transport of macromolecules from any organ proceeds to the blood circulation, we aimed to develop a method that can measure lymphatic function by monitoring the fluorescence in a superficial vein of an interstitially injected tracer. We selected a 40-kDa PEGylated near-infrared dye conjugate, as it showed lymphatic system–specific uptake and extended circulation in blood. Lymphatic transport to blood from subcutaneous tissue required a transit time before signal enhancement was seen in blood followed by a steady rise in signal over time. Increased lymphatic transport was apparent in awake mice compared with those under continuous anesthesia. The methods were validated in K14-VEGFR-3-Fc and K14-VEGF-C transgenic mice with loss and gain of lymphatic function, respectively. Reduced lymphatic transport to blood was also found in aged mice. The technique was also able to measure lymphatic transport from the peritoneal cavity, a location not suitable for optical imaging. The method is a promising, simple approach for assessment of lymphatic function and for monitoring of therapeutic regimens in mouse models of disease and may have potential for clinical translation.

Authors

Steven T. Proulx, Qiaoli Ma, Diana Andina, Jean-Christophe Leroux, Michael Detmar

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Figure 7

Lymphatic transport can be tracked from the peritoneal cavity.

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Lymphatic transport can be tracked from the peritoneal cavity.
(A) Nine-...
(A) Nine-week-old albino female C57BL/6J mice were i.p. injected with 50 μl of 20 μM 40-kDa PEG–IRDye800 conjugate (P40D800) and dynamic imaging of the saphenous vein signal region of interest (ROI) was performed with the mouse in the supine position. (B) Plot showing saphenous vein signal enhancement during anesthetized conditions. Representative of n = 5 mice. (C–E) Images of peritoneal lymphatic outflow 30 minutes after injection of 50 μl of 10 μM 40-kDa PEG–IRDye680 conjugate (P40D680). Left images, Prox1-GFP; middle images, P40D680; right images, merge. Representative of n = 4 mice. (C) Presence of P40D680 in diaphragmatic lymphatic vessels (LVs). Prox1 is expressed also in diaphragmatic muscle. Scale bars: 500 μm. (D) P40D680 in paravertebral LVs (arrows) and thoracic duct (stars). Scale bars: 1 mm. (E) P40D680 in tracheobronchal (right lymph node [LN]) and 2 mediastinal LNs. Scale bars: 1 mm. (F) Representative saphenous vein image at t = 60 minutes after i.p. injection, during which mice were awake and moving normally. (G) Representative saphenous vein image at t = 60 minutes after i.p. injection with mouse under anesthesia. Scale bars: 500 μm. (H) Quantification of the fluorescent signal enhancement at t = 60 minutes comparing mice that were awake with those that were under anesthesia. n = 5 each condition. ***P < 0.001 (2-tailed Student’s t test). Data are the mean ± SD.

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