July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Deciphering subcellular signal sources for optical coherence tomography (OCT) and autofluorescence (AF) imaging of the human retinal pigment epithelium (RPE)
Author Affiliations & Notes
  • Andreas Pollreisz
    Ophthalmology, Medical University Vienna, Vienna, Austria
  • Martina Neschi
    Ophthalmology, Medical University Vienna, Vienna, Austria
  • KENNETH R SLOAN
    University of Alabama at Birmingham, Birmingham, Alabama, United States
  • Tamara Jasmin Mittermüller
    Ophthalmology, Medical University Vienna, Vienna, Austria
  • Dennis M Dacey
    University of Washington, Washington, United States
  • Christine A Curcio
    Ophthalmology, Medical University Vienna, Vienna, Austria
  • Ursula Schmidt-Erfurth
    Ophthalmology, Medical University Vienna, Vienna, Austria
  • Footnotes
    Commercial Relationships   Andreas Pollreisz, None; Martina Neschi, None; KENNETH SLOAN, None; Tamara Mittermüller, None; Dennis Dacey, None; Christine Curcio, Heidelberg Engineering (F), Hoffmann-La Roche (F); Ursula Schmidt-Erfurth, None
  • Footnotes
    Support  Macula Society Research Grant; Macula Foundation; Research to Prevent Blindness, Inc.; EyeSight Foundation of Alabama
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 1574. doi:
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      Andreas Pollreisz, Martina Neschi, KENNETH R SLOAN, Tamara Jasmin Mittermüller, Dennis M Dacey, Christine A Curcio, Ursula Schmidt-Erfurth; Deciphering subcellular signal sources for optical coherence tomography (OCT) and autofluorescence (AF) imaging of the human retinal pigment epithelium (RPE). Invest. Ophthalmol. Vis. Sci. 2019;60(9):1574.

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

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Abstract

Purpose : To quantify the total number, dimensions, and spatial distribution of signal generating organelles in the cell body (CB) and apical processes (AP) of the human RPE using 3D connectomics technology.

Methods : A whole globe of a 28-year-old male Caucasian organ donor with an unremarkable macula was processed via rapid organ recovery and perifoveal epoxy-embedded RPE imaged by serial block-face scanning electron microscopy. The image stack was annotated manually by expert readers using TrakEM (ImageJ).

Results : The 3D dataset showed exquisite ultrastructural preservation allowing the reconstruction of entire RPE cells including AP and CB. In 17 completely reconstructed RPE cells (including 2 binucleate) melanosomes (M) and lipofuscin granules (L) were identified in AP and tracked to their respective CB. The mononucleate RPE cells comprised a mean number of 65±24 (standard deviation) M and 12±6 L. Binucleate RPE cells contained 131±28 M and 26±1 L per cell, with each organelle filling the entire cross-section of AP. In a single representative RPE cell, the mean length, width and volume for M and L were 2639±588 nm, 619±99 nm, 0.82±0.3 µm3 and 2997±1237 nm, 610±150 nm, 0.87±0.33 µm3, respectively. RPE cell bodies had a mean volume of 2416±263 µm3 per cell and contained mitochondria (Mi), L, M and melanolipofuscin (ML) granules. The mean number of Mi per RPE CB was 734±170. The mean volume of 767 Mi in a representative RPE CB was 0.24±0.12 µm3 comprising 8% of CB volume. The combined number of L, M and melanolipofuscin (ML) granules per RPE cell body was 681±153. Accounting for all measured reflective granules in AP and CB revealed 85% coverage of the retinal image plane per RPE cell, implying that posterior structures are 85% shadowed by healthy RPE.

Conclusions : A 3D connectomics approach allowed the complete volumetric reconstruction of entire human RPE cells including organelles serving as signal sources for OCT (all) and AF (L, ML) imaging. Our results revealed that each RPE cell has hundreds of Mi, L, LF, and M with a surface coverage of 85% contributing to reflectivity via Mie scattering. These results will impact clinical interpretation of OCT and AF imaging and theories of macular disease pathophysiology.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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