Abstract
Purpose :
The aim of this study was to reveal the effect of mesalazine on proinflammatory cytokines in the PBMCs of patients with HLA-B27 associated acute anterior uveitis (AAU), and whether the mechanism of 5-ASA action is dependent of the HLA-B27 status of patients (HLA-B27+ vs. HLA-B27-).
Methods :
Human PBMCs were isolated from healthy donors and patients with AAU (HLA-B27+ (N=16) and HLA-B27- (N=6)) using Biocoll gradient centrifugation. After prestimulation with Sulfasalazine (10µM and 100µM) and mesalazine (mesalazine 5-ASA) (100µM and 500µM) for 30 minutes, cells were stimulated with the TLR 9 ligand LPS and the TLR 8 ligand resiquimod (R848) (10 µg/ml) for additional 2.5h. To investigate whether endoplasmatic reticulum (ER) stress is involved in the response to 5-ASA treatment we induced ER stress using (4µM) thapsigargin (Tsg). To determine gene expression of different proinflammatory cytokines (IL-6, IL-1, TNFα, IFNγ, IL17A and IL23A) quantitative real-time-PCR has been used. Human IL-6 and IL-1b secretion was measured in cell-free supernatants using ELISA.
Results :
In a first step the impact of sulfasalazine on IL-6 and IL-1β had been studied non-uveitic persons. No suppression on mRNA and protein level had been observed. After using mesalazine (a metabolic derivate of sulfasalazine) on the cells of healthy donors mesalazine altered IL-6 and IL-1β on both levels after LPS treatment, but not after R848 treatment. Treating PBMCs derived from patients with AAU, HLA-B27 positive and HLA-B27-negative showed suppression of IL-6 on mRNA and protein level. No suppression was detectable for IL-1β and TNFα in both patients cohorts. In a second step, we observed suppression of all ER stress related cytokines (IL-6, TNF, IFNγ, IL17A and IL23A) in the HLA-B27 positive and in the negative patients after using mesalazine
Conclusions :
After 5-ASA treatment in healthy donors we observed a suppression of the proinflammatory cytokines IL-6 and IL-1 on mRNA and protein level. In patients with HLA-B27 associated AAU, suppression of IL-6 has been observed on mRNA and protein level and that inhibition was independent of HLA-B27 status of patients. ER stress associated cytokines such as IFNγ, IL17A and IL23A had been suppressed by 5-ASA independent of HLA-B27 status of the patients. In conclusion 5-ASA suppressed UPR associated cytokine secretion in human PBMCs independently whether patients were HLA-B27 positive or negative.
This is a 2020 ARVO Annual Meeting abstract.