Adult human RPE has greater susceptibility to IL17A-induced degeneration than fetal human RPE

Abstract

Purpose: Age-related macular degeneration (AMD) results from alteration of the retinal pigment epithelium (RPE) and subsequent atrophy of photoreceptors in the macula. Elevated expression of IL17A transcripts has been found in human AMD maculae in addition to reports of elevated IL17A in AMD patient sera. Most recently, hypomethylation of the IL17 receptor C (IL17RC) promoter has been linked to AMD. While IL17A is known to generate immune response, its role in RPE degeneration remains largely unclear. Here, we compare the effect of IL17A on primary adult and fetal human RPE.

Methods: Primary adult and fetal human RPE cells were serum-starved for 24h and then treated for 48h with recombinant IL17A. Expression of IL17 receptor A (IL17RA) and IL17RC was detected using quantitative RT-PCR. MTT assays were performed to evaluate RPE cell viability in response to varying doses of IL17A. Transmission electron microscopy (TEM) was used to compare untreated and IL17A-treated cells for ultrastructural changes. Immunohistochemical detection of capase-9 activation was used to evaluate apoptosis as well as caspase-1 for inflammasome activity.

Results: IL17A-treated adult and fetal RPE cells showed elevated expression of IL17RA and IL17RC transcripts in comparison to untreated cells. Adult RPE had greater sensitivity to IL17A than fetal RPE. MTT assays showed dosage-independent decrease in cell viability of IL17A-treated adult and fetal RPE cells, with no significant difference in response between these two cell lines. At baseline, TEM revealed more cytoplasmic keratin filaments in adult RPE than in fetal RPE and after treatment, adult RPE showed increased vacuolization and mitochondrial damage, in addition to greater numbers of autophagosomes and lysosomes than fetal RPE. Immunohistochemistry detected increased caspase-9 and caspase-1 activation in adult RPE as compared to fetal RPE.

Conclusions: IL17A is capable of inducing RPE degeneration in both adult and fetal cell lines. Adult RPE appear to be more sensitive to IL17A treatment than fetal RPE, suggesting adult RPE may be more inclined to homeostatic imbalance than fetal RPE and thus, more susceptible to IL17A-induced degeneration.

Keywords: 412 age-related macular degeneration • 490 cytokines/chemokines • 426 apoptosis/cell death