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Updated: Jul 8, 2026
Differentiation, Maintenance, and Analysis of Human Retinal Pigment Epithelium Cells: A Disease-in-a-dish Model for BEST1 Mutations
Published on: August 24, 2018
Distinct P2Y receptor subtypes regulate calcium signaling in human retinal pigment epithelial cells
Victoria E Tovell1, Julie Sanderson
1School of Chemical Sciences and Pharmacy, University of East Anglia, Norwich, United Kingdom.
Purpose:
Nucleotide signaling plays a role in retinal pigment epithelial (RPE) function, and receptors for nucleotides are potential therapeutic targets for various ocular diseases. The purpose of this study was to investigate the expression of P2Y receptor subtypes in native and cultured human RPE cells.
Methods:
Intracellular Ca(2+) levels were monitored using real-time fluorescence imaging in cultured human RPE cells loaded with Fura-2. Expression of P2Y receptors in native and cultured RPE cells was determined by quantitative RT-PCR and Western blot analysis.
Results:
Adenosine triphosphate (ATP), uridine triphosphate (UTP), adenosine diphosphate (ADP), 2-methylthio ATP (2MeSATP), and uridine diphosphate (UDP) produced concentration-related increases in [Ca(2+)](i) in cultured RPE cells. However, differences between the magnitude and shape of agonist responses were observed. ATP and UTP showed similar response characteristics, including a distinct Ca(2+) influx component. ATP and UTP were equipotent (EC(50), 6 muM) and maximum responses were equivalent, suggesting activation of a P2Y(2) receptor. Maximal responses to ADP and 2MeSATP were equivalent with EC(50)s of 1 muM and 0.3 muM. The P2Y(1) antagonist MRS 2179 (10 muM) inhibited these responses, confirming functional expression of P2Y(1) receptors. The presence of a response to UDP suggested P2Y(6) expression. There was no influx component to P2Y(1)- and P2Y(6)-mediated responses. mRNA for P2Y(1), P2Y(2,) P2Y(4), and P2Y(6) receptor subtypes was found in cultured RPE cells, and for P2Y(1), P2Y(2,) P2Y(4,) P2Y(6), and P2Y(12) it was found in native RPE cells. Expression of P2Y(1), P2Y(2), and P2Y(6) protein was found in native and cultured RPE cells.
Conclusions:
These data define the expression profile of P2Y receptors in human RPE and show that different P2Y subtypes control distinct calcium responses in these cells.
Insights
This study investigated P2Y receptor expression in human retinal pigment epithelial (RPE) cells. We found specific P2Y subtypes are expressed and functionally active, influencing calcium signaling in RPE cells.
Area of Science:
- Ophthalmology
- Cell Biology
- Neuroscience
Background:
- Nucleotide signaling is crucial for retinal pigment epithelial (RPE) cell function.
- P2Y receptors are potential therapeutic targets for ocular diseases.
Purpose of the Study:
- To investigate the expression profile of P2Y receptor subtypes in native and cultured human RPE cells.
- To determine the functional roles of these receptors in RPE calcium signaling.
Main Methods:
- Real-time fluorescence imaging of intracellular calcium levels in cultured RPE cells.
- Quantitative RT-PCR and Western blot analysis to assess P2Y receptor mRNA and protein expression.
Main Results:
- Cultured RPE cells responded to various nucleotides (ATP, UTP, ADP, 2MeSATP, UDP), indicating functional P2Y receptor expression.
- Evidence suggests the presence of P2Y(1), P2Y(2), and P2Y(6) receptors, with distinct calcium response characteristics.
- mRNA for P2Y(1), P2Y(2), P2Y(4), P2Y(6), and P2Y(12) subtypes was detected in RPE cells.
Conclusions:
- This study defines the P2Y receptor expression profile in human RPE cells.
- Different P2Y receptor subtypes mediate distinct calcium responses in RPE cells, highlighting their complex role in ocular function.
