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In Vitro Method to Study Sex-Based Differences in Conjunctival Goblet Cells

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In Vitro Method to Study Sex-Based Differences in Conjunctival Goblet Cells.

Jeffrey A Bair¹, Darlene A Dartt¹, Menglu Yang²

¹Department of Ophthalmology Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School.

Journal of Visualized Experiments : Jove

|August 14, 2023

Summary

Researchers developed a method to remove exogenous hormones from cell culture media, crucial for studying sex-based differences in conjunctival goblet cells (CGCs). This technique ensures CGCs maintain normal function, vital for dry eye disease research.

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Area of Science:

Ophthalmology
Cell Biology
Endocrinology

Background:

Dry eye disease is multifactorial, impacting ocular surface health, with higher prevalence in women.
Disruption of mucin secreted by conjunctival goblet cells (CGCs) contributes to ocular surface diseases.
Exogenous sex hormones in culture media can confound in vitro studies of sex-based differences in CGCs.

Purpose of the Study:

To describe a method for minimizing exogenous hormones in CGC cultures.
To maintain the physiological function of CGCs during hormone elimination.
To enable consistent in vitro study of sex-based differences in CGCs.

Main Methods:

Conjunctival goblet cells (CGCs) from human donors were cultured.
Cells were transferred to phenol-red-free RPMI medium with 1% BSA 48 hours before experiments.

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Intracellular calcium ([Ca2+]i) response to carbachol stimulation was measured using fura 2/acetoxymethyl (AM) microscopy.

Main Results:

CGCs maintained normal function in phenol-red-free medium after 48 hours.
No significant difference in intracellular calcium ([Ca2+]i) response was observed compared to complete medium.
The method effectively eliminates exogenous hormones without altering CGC physiological function.

Conclusions:

Phenol-red-free RPMI medium with 1% BSA is recommended for studying sex-based differences in CGCs.
This method removes exogenous hormones while preserving normal CGC function.
Essential for accurate research into sex-based variations in ocular surface health and disease.