Electron microscopic localization of transthyretin in cultured and native HRPE

R. K. Getz, Brian Kennedy, N. J. Mangini

Research output: Contribution to journalArticle

Abstract

Purpose. Transthyretin (TTR) is a 55 kD tetrameric protein which binds thyroxine (T4) as well as the complex formed by retinol and retinol binding protein. TTR could play a role in the intraocular cycling of vitamin A and thyroxine. This study was performed to determine the intracellular localization of TTR in cultured and native human retinal pigment epithelial cells (HRPE). Methods. Immunoreactivity to TTR was localized at the electron microscopic level in fixed monolayer cultures and native tissue isolated from adult human donor tissue, using monoclonal antibodies (mAbs) (6E5-2 and 14D1-1) against TTR in a post-embedding immunogold technique. Tissues were embedded with Lowicryl K4M, and labeling was performed with 20 nm gold-tagged goat anti-mouse IgG. Results. Immunoreactivity was observed over the nucleus and mitochondria. In the cytoplasm, reaction product was detected in a non-vesicle bound, random distribution. Immunoreactivity was also detected on the plasma membrane. It was observed on both apical and basolateral surfaces, but was most abundant apically. The labeling pattern was the same in cultured and native preparations and for both mAbs. Mouse IgG, used as a negative control, did not exhibit labeling. Conclusions. The subcellular localization of TTR in HRPE cells grown in culture parallels that of native HRPE cells, and suggests an intracellular function for TTR, possibly in association with T4; in addition to its previously hypothesized role in retinol transport.

Original languageEnglish (US)
JournalInvestigative Ophthalmology and Visual Science
Volume37
Issue number3
StatePublished - Feb 15 1996

Fingerprint

Prealbumin
Retinal Pigments
Epithelial Cells
Electrons
Vitamin A
Thyroxine
Monoclonal Antibodies
Retinol-Binding Proteins
Population Groups
Goats
Gold
Mitochondria
Cytoplasm
Immunoglobulin G
Immunohistochemistry
Cell Membrane
Tissue Donors

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Electron microscopic localization of transthyretin in cultured and native HRPE. / Getz, R. K.; Kennedy, Brian; Mangini, N. J.

In: Investigative Ophthalmology and Visual Science, Vol. 37, No. 3, 15.02.1996.

Research output: Contribution to journalArticle

@article{54af8c2f97434a8ba33d028f6e0261b5,
title = "Electron microscopic localization of transthyretin in cultured and native HRPE",
abstract = "Purpose. Transthyretin (TTR) is a 55 kD tetrameric protein which binds thyroxine (T4) as well as the complex formed by retinol and retinol binding protein. TTR could play a role in the intraocular cycling of vitamin A and thyroxine. This study was performed to determine the intracellular localization of TTR in cultured and native human retinal pigment epithelial cells (HRPE). Methods. Immunoreactivity to TTR was localized at the electron microscopic level in fixed monolayer cultures and native tissue isolated from adult human donor tissue, using monoclonal antibodies (mAbs) (6E5-2 and 14D1-1) against TTR in a post-embedding immunogold technique. Tissues were embedded with Lowicryl K4M, and labeling was performed with 20 nm gold-tagged goat anti-mouse IgG. Results. Immunoreactivity was observed over the nucleus and mitochondria. In the cytoplasm, reaction product was detected in a non-vesicle bound, random distribution. Immunoreactivity was also detected on the plasma membrane. It was observed on both apical and basolateral surfaces, but was most abundant apically. The labeling pattern was the same in cultured and native preparations and for both mAbs. Mouse IgG, used as a negative control, did not exhibit labeling. Conclusions. The subcellular localization of TTR in HRPE cells grown in culture parallels that of native HRPE cells, and suggests an intracellular function for TTR, possibly in association with T4; in addition to its previously hypothesized role in retinol transport.",
author = "Getz, {R. K.} and Brian Kennedy and Mangini, {N. J.}",
year = "1996",
month = "2",
day = "15",
language = "English (US)",
volume = "37",
journal = "Investigative Ophthalmology and Visual Science",
issn = "0146-0404",
publisher = "Association for Research in Vision and Ophthalmology Inc.",
number = "3",

}

TY - JOUR

T1 - Electron microscopic localization of transthyretin in cultured and native HRPE

AU - Getz, R. K.

AU - Kennedy, Brian

AU - Mangini, N. J.

PY - 1996/2/15

Y1 - 1996/2/15

N2 - Purpose. Transthyretin (TTR) is a 55 kD tetrameric protein which binds thyroxine (T4) as well as the complex formed by retinol and retinol binding protein. TTR could play a role in the intraocular cycling of vitamin A and thyroxine. This study was performed to determine the intracellular localization of TTR in cultured and native human retinal pigment epithelial cells (HRPE). Methods. Immunoreactivity to TTR was localized at the electron microscopic level in fixed monolayer cultures and native tissue isolated from adult human donor tissue, using monoclonal antibodies (mAbs) (6E5-2 and 14D1-1) against TTR in a post-embedding immunogold technique. Tissues were embedded with Lowicryl K4M, and labeling was performed with 20 nm gold-tagged goat anti-mouse IgG. Results. Immunoreactivity was observed over the nucleus and mitochondria. In the cytoplasm, reaction product was detected in a non-vesicle bound, random distribution. Immunoreactivity was also detected on the plasma membrane. It was observed on both apical and basolateral surfaces, but was most abundant apically. The labeling pattern was the same in cultured and native preparations and for both mAbs. Mouse IgG, used as a negative control, did not exhibit labeling. Conclusions. The subcellular localization of TTR in HRPE cells grown in culture parallels that of native HRPE cells, and suggests an intracellular function for TTR, possibly in association with T4; in addition to its previously hypothesized role in retinol transport.

AB - Purpose. Transthyretin (TTR) is a 55 kD tetrameric protein which binds thyroxine (T4) as well as the complex formed by retinol and retinol binding protein. TTR could play a role in the intraocular cycling of vitamin A and thyroxine. This study was performed to determine the intracellular localization of TTR in cultured and native human retinal pigment epithelial cells (HRPE). Methods. Immunoreactivity to TTR was localized at the electron microscopic level in fixed monolayer cultures and native tissue isolated from adult human donor tissue, using monoclonal antibodies (mAbs) (6E5-2 and 14D1-1) against TTR in a post-embedding immunogold technique. Tissues were embedded with Lowicryl K4M, and labeling was performed with 20 nm gold-tagged goat anti-mouse IgG. Results. Immunoreactivity was observed over the nucleus and mitochondria. In the cytoplasm, reaction product was detected in a non-vesicle bound, random distribution. Immunoreactivity was also detected on the plasma membrane. It was observed on both apical and basolateral surfaces, but was most abundant apically. The labeling pattern was the same in cultured and native preparations and for both mAbs. Mouse IgG, used as a negative control, did not exhibit labeling. Conclusions. The subcellular localization of TTR in HRPE cells grown in culture parallels that of native HRPE cells, and suggests an intracellular function for TTR, possibly in association with T4; in addition to its previously hypothesized role in retinol transport.

UR - http://www.scopus.com/inward/record.url?scp=33750183942&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33750183942&partnerID=8YFLogxK

M3 - Article

VL - 37

JO - Investigative Ophthalmology and Visual Science

JF - Investigative Ophthalmology and Visual Science

SN - 0146-0404

IS - 3

ER -