In vivo imaging of the human eye using a 2-photon-excited fluorescence scanning laser ophthalmoscope

Jakub Boguslawski; Grazyna Palczewska; Slawomir Tomczewski; Jadwiga Milkiewicz; Piotr Kasprzycki; Dorota Stachowiak; Katarzyna Komar; Marcin J Marzejon; Bartosz L Sikorski; Arkadiusz Hudzikowski; Aleksander Głuszek; Zbigniew Łaszczych; Karol Karnowski; Grzegorz Soboń; Krzysztof Palczewski; Maciej Wojtkowski

doi:10.1172/JCI154218

In vivo imaging of the human eye using a 2-photon-excited fluorescence scanning laser ophthalmoscope

J Clin Invest. 2022 Jan 18;132(2):e154218. doi: 10.1172/JCI154218.

Authors

Jakub Boguslawski^{1

2

3}, Grazyna Palczewska^{1

2

4

5}, Slawomir Tomczewski^{1

2}, Jadwiga Milkiewicz^{1

2}, Piotr Kasprzycki^{1

2}, Dorota Stachowiak³, Katarzyna Komar^{1

2

6}, Marcin J Marzejon^{1

2

7}, Bartosz L Sikorski^{8

9}, Arkadiusz Hudzikowski³, Aleksander Głuszek³, Zbigniew Łaszczych³, Karol Karnowski^{1

2}, Grzegorz Soboń³, Krzysztof Palczewski^{5

10

11

12}, Maciej Wojtkowski^{1

2

6}

Affiliations

¹ International Center for Translational Eye Research and.
² Department of Physical Chemistry of Biological Systems, Institute of Physical Chemistry, Polish Academy of Sciences, Warsaw, Poland.
³ Laser and Fiber Electronics Group, Faculty of Electronics, Photonics and Microsystems, Wrocław University of Science and Technology, Wrocław, Poland.
⁴ Department of Medical Devices, Polgenix, Inc., Cleveland, Ohio, USA.
⁵ Department of Ophthalmology, Gavin Herbert Eye Institute, University of California, Irvine, California, USA.
⁶ Faculty of Physics, Astronomy and Informatics, Nicolaus Copernicus University, Torun, Poland.
⁷ Department of Metrology and Optoelectronics, Faculty of Electronics, Telecommunications and Informatics, Gdansk University of Technology, Gdansk, Poland.
⁸ Department of Ophthalmology, Nicolaus Copernicus University, Bydgoszcz, Poland.
⁹ Oculomedica Eye Research & Development Center, Bydgoszcz, Poland.
¹⁰ Department of Physiology & Biophysics, School of Medicine.
¹¹ Department of Chemistry, and.
¹² Department of Molecular Biology & Biochemistry, University of California, Irvine, California, USA.

Abstract

BackgroundNoninvasive assessment of metabolic processes that sustain regeneration of human retinal visual pigments (visual cycle) is essential to improve ophthalmic diagnostics and to accelerate development of new treatments to counter retinal diseases. Fluorescent vitamin A derivatives, which are the chemical intermediates of these processes, are highly sensitive to UV light; thus, safe analyses of these processes in humans are currently beyond the reach of even the most modern ocular imaging modalities.MethodsWe present a compact, 2-photon-excited fluorescence scanning laser ophthalmoscope and spectrally resolved images of the human retina based on 2-photon excitation (TPE) with near-infrared light. A custom Er:fiber laser with integrated pulse selection, along with intelligent postprocessing of data, enables excitation with low laser power and precise measurement of weak signals.ResultsWe demonstrate spectrally resolved TPE fundus images of human subjects. Comparison of TPE data between human and mouse models of retinal diseases revealed similarity with mouse models that rapidly accumulate bisretinoid condensation products. Thus, visual cycle intermediates and toxic byproducts of this metabolic pathway can be measured and quantified by TPE imaging.ConclusionOur work establishes a TPE instrument and measurement method for noninvasive metabolic assessment of the human retina. This approach opens the possibility for monitoring eye diseases in the earliest stages before structural damage to the retina occurs.FundingNIH, Research to Prevent Blindness, Foundation for Polish Science, European Regional Development Fund, Polish National Agency for Academic Exchange, and Polish Ministry of Science and Higher Education.

Keywords: Medical devices; Ophthalmology.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Disease Models, Animal
Female
Humans
Male
Mice
Mice, Knockout
Middle Aged
Ophthalmoscopes*
Optical Imaging*
Retina* / diagnostic imaging
Retina* / metabolism
Retinal Diseases* / diagnostic imaging
Retinal Diseases* / genetics
Retinal Diseases* / metabolism

Grants and funding

The International Centre for Translational Eye Research (MAB/2019/12) project is carried out within the International Research Agendas Programme of the Foundation for Polish Science co-financed by the European Union under the European Regional Development Fund. The 2x2 PhotonVis project no. POIR.04.04.00-00-3D47/16-00 is carried out within the TEAM TECH Programme of the Foundation for Polish Science co-financed by the European Union under the European Regional Development Fund. This work was partially supported by the Foundation for Polish Science within the First TEAM program co-financed by the European Union under the European Regional Development Fund (First TEAM/2017-4/39). The authors also acknowledge support from an RPB unrestricted grant to the Department of Ophthalmology, University of California, Irvine. K.K. acknowledges National Science Centre, grant 2016/23/B/ST2/00752. P.K. and KK were supported by the Polish National Agency for Academic Exchange (PPN/PPO/2018/1/00082) and Polish Ministry of Science and Higher Education (2016-2019 int.).