Understanding Inflammation-associated Ophthalmic Pathologies: A Novel 3D Co-culture Model of Monocyte-myofibroblast Immunomodulation

ABSTRACT

Purpose

Inflammation is associated with, and may be causal of, a variety of ophthalmic pathologies. These pathologies are currently difficult to model in vitro because they involve complex interactions between the innate immune system, stromal cells, and other cells that normally maintain ocular tissue homeostasis. Using transscleral drainage channel fibrosis after glaucoma surgery as an example of inflammation-associated ocular fibrosis, we have assessed a simple but novel 3D cell culture system designed to reveal the immunomodulatory impacts of ocular connective tissue cells on monocytes, a major cellular component of the circulating immune system.

Methods

Primary human Tenon’s capsule fibroblasts derived from five unrelated patients were activated into myofibroblasts in 3D collagen matrices under isometric tension, with and without exposure to an inflammatory cytokine-enhanced milieu, and co-cultured with an immortalized human monocyte cell line (THP-1 cells). Quantitative PCR analyses were performed on 8 candidate genes to assess the impacts of inflammatory cytokines on the myofibroblasts and the monocytes in mono-cultures and compared to cells in co-culture to clearly distinguish any co-culture-induced impacts on gene expression.

Results

Our data indicate that both Tenon’s capsule myofibroblasts in 3D mono-culture and THP-1 monocytes in suspension mono-culture were responsive to inflammatory cytokine stimuli. Co-culture with Tenon’s capsule myofibroblasts significantly modulated the gene expression responses of THP-1 monocytes to inflammatory cytokine stimulation, indicative of an immunomodulatory “feedback” system between these cell types.

Conclusion

Our findings provide proof of principle for the use of simple 3D co-culture systems as a means to enhance our understanding of ocular stromal cell interactions with cells of the innate immune system and to provide more informative in vitro models of inflammation-associated ophthalmic pathologies.

KEYWORDS:

• Inflammation
• Myofibroblasts
• Monocytes
• Glaucoma
• Fibrosis
• Co-culture
• 3d model

Acknowledgments

I acknowledge the students and staff of the O’Gorman and Hutnik laboratories for their support, communication, and mentorship for the duration of this project.

Declaration of interest

The authors report no conflict of interest. The authors report funding from the Schulich School of Medicine and Dentistry, Western University. The content of this submission has not been published or submitted for publication elsewhere. All authors have contributed significantly and agree with the content of this manuscript.

Additional information

Funding

This work was supported by the Western University.