Expression of Cytokines in Porcine Iris, Retina and Choroidal Tissues Stimulated by Microbe-associated Molecular Patterns

ABSTRACT

Purpose

The innate immune system is strongly implicated in the pathogenesis of uveitis. This study was designed to clarify the responses of the innate immune system in uveal tissues.

Materials and Methods

We utilized quantitative, real-time RT-PCR to measure mRNA of innate immune system receptors from porcine iris, choroid, and retina tissues. We used RT-PCR for cytokines to evaluate the responses of these tissues to specific ligands or extracts of whole bacteria that activate the innate immune system. We used ELISA for IL-6 on selected choroidal supernatants to confirm that the mRNA measurement correlated with protein levels.

Results

In each of the studied tissues, we detected the expression of important receptors belonging to the innate immune system including dectin-1, TLR4, TLR8, and NOD2. Relative mRNA expression was generally lower in the retina compared to iris or choroid. All three tissues demonstrated upregulation of cytokine mRNA in response to a range of ligands that activate the innate immune system. The measurement of IL-6 protein was consistent with results based on mRNA. Notably, the expression of mRNA for IL-23 was more pronounced than IL-12 in all three tissues after stimulation with various innate immune system ligands.

Conclusions

These data provide evidence of a potent innate immune response intrinsic to uveal tissues. Specific innate immune system ligands as well as bacterial extracts enhanced the production of several inflammatory cytokines. Furthermore, the observation of higher upregulation of IL-23 mRNA, compared to IL-12 in response to innate immune stimuli, suggested that a local TH17 response might be more robust than a local TH1 response in uveal tissues. Our results expand the understanding as to how the innate immune system may contribute to uveitis.

KEYWORDS:

• Iris
• retina
• choroid
• pattern recognition receptors
• innate immunity
• microbe-associated molecular patterns
• cytokines

Acknowledgments

This study was supported by the biomedical research institute fund (GNUHBRIF-2018-0002) by the Gyeongsang National University Hospital and the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (grant numbers NRF- 2019R1F1A1061541). We are also grateful for the support of the Spondylitis Association of America, the William and Mary Bauman Family Foundation, the Stan and Madelle Rosenfeld Family Trust, and the Grandmaison Fund for Autoimmunity Research; and unrestricted departmental funding to the Casey Eye Institute from Research to Prevent Blindness (New York, NY). Melanie Alvarado and Naman Jain provided technical support.

The following reagent was obtained through BEI Resources, NIAID, NIH as part of the Human Microbiome Project: Parabacteroides distasonis, Strain 31_2 (Deposited as Porphyromonas sp., Strain 31_2), HM-169.

Declaration of interests

Y.S. Han, None; E. Rivera-Grana, None; J.T. Rosenbaum, Commercial Relationship: Consultant for Abbvie, Regeneron, Gilead, UCB, Eyevensys, Novartis, Janssen, Santen, Horizon, and Roche unrelated to this work. Royalties from UpToDate and research funds from Pfizer; M. Schleisman, None; S. Davin, None; T.M. Martin, None; A.B. Furst, None; M. Asquith, None.