Characterization of Rat Meibomian Gland Ion and Fluid Transport

Dongfang Yu; Richard M Davis; Megumi Aita; Kimberlie A Burns; Phillip W Clapp; Rodney C Gilmore; Michael Chua; Wanda K O'Neal; Richard Schlegel; Scott H Randell; Richard C Boucher

doi:10.1167/IOVS.15-17945

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Characterization of Rat Meibomian Gland Ion and Fluid Transport

Journal article

Open access

Peer reviewed

Characterization of Rat Meibomian Gland Ion and Fluid Transport

Dongfang Yu, Richard M Davis, Megumi Aita, Kimberlie A Burns, Phillip W Clapp, Rodney C Gilmore, Michael Chua, Wanda K O'Neal, Richard Schlegel, Scott H Randell, …

Investigative ophthalmology & visual science, Vol.57(4), pp.2328-2343

04/01/2016

DOI: 10.1167/IOVS.15-17945

PMCID: PMC4855829

PMID: 27127933

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https://doi.org/10.1167/IOVS.15-17945View

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Abstract

We establish novel primary rat meibomian gland (MG) cell culture systems and explore the ion transport activities of the rat MG. Freshly excised rat MG tissues were characterized as follows: (1) mRNA expression of selected epithelial ion channels/transporters were measured by RT-PCR, (2) localization of epithelial sodium channel (ENaC) mRNAs was performed by in situ hybridization, and (3) protein expression and localization of βENaC, the Na+/K+/Cl- cotransporter (NKCC), and the Na+/K+ ATPase were evaluated by immunofluorescence. Primary isolated rat MG cells were cocultured with 3T3 feeder cells and a Rho-associated kinase (ROCK) inhibitor (Y-27632) for expansion. Passaged rat MG cells were cultured as planar sheets under air-liquid interface (ALI) conditions for gene expression and electrophysiologic studies. Passaged rat MG cells also were cultured in matrigel matrices to form spheroids, which were examined ultrastructurally by transmission electron microscopy (TEM) and functionally using swelling assays. Expression of multiple ion channel/transporter genes was detected in rat MG tissues. β-ENaC mRNA and protein were localized more to MG peripheral acinar cells than central acinar cells or ductular epithelial cells. Electrophysiologic studies of rat MG cell planar cultures demonstrated functional sodium, chloride, and potassium channels, and cotransporters activities. Transmission electron microscopic analyses of rat MG spheroids revealed highly differentiated MG cells with abundant lysosomal lamellar bodies. Rat MG spheroids culture-based measurements demonstrated active volume regulation by ion channels. This study demonstrates the presence and function of ion channels and volume transport by rat MG. Two novel primary MG cell culture models that may be useful for MG research were established.

Amides - pharmacology

Microscopy, Electron, Transmission

Sodium-Potassium-Exchanging ATPase - physiology

Coculture Techniques

Cells, Cultured

Rats

Male

Ion Channels - physiology

Rats, Sprague-Dawley

Meibomian Glands - physiology

rho-Associated Kinases - antagonists & inhibitors

Animals

In Situ Hybridization

Meibomian Glands - metabolism

Sodium-Potassium-Chloride Symporters - physiology

Fluorescent Antibody Technique

Mice

Pyridines - pharmacology

Ion Transport - physiology

3T3 Cells - physiology

Meibomian Glands - cytology

Details

Title: Subtitle: Characterization of Rat Meibomian Gland Ion and Fluid Transport
Creators: Dongfang Yu - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Richard M Davis - Department of Ophthalmology, The University of North Carolina, Chapel Hill, North Carolina, United States
Megumi Aita - Neuroscience Center, The University of North Carolina, Chapel Hill, North Carolina, United States
Kimberlie A Burns - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Phillip W Clapp - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Rodney C Gilmore - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Michael Chua - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Wanda K O'Neal - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Richard Schlegel - Department of Pathology, Georgetown University Medical School, Washington District of Columbia, United States
Scott H Randell - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Richard C Boucher - Marsico Lung Institute/UNC Cystic Fibrosis Research Center School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States
Resource Type: Journal article
Publication Details: Investigative ophthalmology & visual science, Vol.57(4), pp.2328-2343
Publisher: United States
DOI: 10.1167/IOVS.15-17945
PMID: 27127933
PMCID: PMC4855829
ISSN: 0146-0404
eISSN: 1552-5783
Grant note: P50 HL107168 / NHLBI NIH HHS P01HL108808 / NHLBI NIH HHS P01 HL110873 / NHLBI NIH HHS P01 HL108808 / NHLBI NIH HHS P01HL110873 / NHLBI NIH HHS P30DK065988 / NIDDK NIH HHS P50HL107168 / NHLBI NIH HHS P30 DK065988 / NIDDK NIH HHS
Language: English
Date published: 04/01/2016
Academic Unit: Periodontics
Record Identifier: 9984065719002771

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