April 2014
Volume 55, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2014
Imaging of Retinal Vascular Disease Using Hypoxia-Sensitive Contrast Agents
Author Affiliations & Notes
  • Stephanie M. Evans
    Ophthalmology and Visual Sciences, Vanderbilt Eye Institute, Nashville, TN
  • Ashwath Jayagopal
    Ophthalmology and Visual Sciences, Vanderbilt Eye Institute, Nashville, TN
    Molecular Physiology and Biophysics, Vanderbilt University Medical Center, Nashville, TN
  • Footnotes
    Commercial Relationships Stephanie Evans, None; Ashwath Jayagopal, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 5873. doi:
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      Stephanie M. Evans, Ashwath Jayagopal; Imaging of Retinal Vascular Disease Using Hypoxia-Sensitive Contrast Agents. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5873.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract
 
Purpose
 

Retinal vascular diseases, including diabetic retinopathy and age related macular degeneration, are often associated with retinal hypoxia. Therefore, the ability to image hypoxic retinal tissue in vivo would be beneficial for improved clinical management of these diseases. For this purpose, a hypoxia-sensitive fluorescent contrast agent was developed and characterized for imaging of hypoxia in retinal tissue using established cell culture and animal models of retinal vascular disease.

 
Methods
 

Fluorescently-labeled, hypoxia-sensitive contrast agents were synthesized, purified chromatographically, and characterized by mass spectrometric and nuclear magnetic resonance analyses. To evaluate the utility of this contrast agent for imaging hypoxia, in vitro assays using Human Retinal Microvascular Endothelial Cells (HRMEC) and in vivo studies using mice with oxygen-induced retinopathy (OIR) or laser-induced choroidal neovascularization (LCNV) were performed to determine the hypoxia-associated sensitivity and specificity of this contrast agent. Fluorimetric and flow cytometric assays were performed on normoxia- or hypoxia-conditioned HRMEC exposed to contrast agents to measure contrast agent binding and uptake. OIR and LCNV animal models were intravenously or intraocularly-injected with the contrast agents and analyzed by in vivo and ex vivo retinal fluorescence imaging to determine specificity and sensitivity of the contrast agent. Pimonidazole hydrocholoride immunostaining was utilized to confirm specificity of the contrast agent for hypoxic tissue.

 
Results
 

HRMEC conditioned under hypoxia for varying durations of time up to 24 hrs. exhibited dose-dependent fluorescence enhancement due to hypoxia-selective uptake of the contrast agent. In animal models, regions of tissue hypoxia staining positive for pimonidazole hydrochloride were also colocalized with contrast agent uptake, as indicated by in vivo and ex vivo imaging. Contrast agent accumulation in hypoxic tissue was detectable within 2 hrs. post-injection.

 
Conclusions
 

These studies support the feasibility of imaging hypoxic tissue in vivo using targeted contrast agents in conjunction with readily available retinal fluorescence imaging equipment. Hypoxia-sensitive contrast agents, if clinically translated, may be useful for early detection of retinal vascular diseases and monitoring of therapeutic response in patients.

 
Keywords: 548 hypoxia • 552 imaging methods (CT, FA, ICG, MRI, OCT, RTA, SLO, ultrasound) • 688 retina  
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