Individual components of a nutraceutical formula synergistically contribute to the effect on COX-2 and iNos activity and expression

Abstract

Purpose : Anti-VEGF agents currently available for the treatment of Diabetic Macular Edema and Wet Age-Related Macular Degeneration do not possess a direct effect on inflammation. Here we set out to investigate whether a nutritional supplement may aid the activity of anti-VEGF drugs by providing anti-inflammatory support.

Methods : A formula codenamed AVS (SIFI S.p.A.) (1.9 mg/ml) and its individual components (astaxanthin, piperine, boswellic acid, bromelain, coenzyme Q10, copper, curcumin, lutein, anthocyanin, resveratrol, safranal, salicin, zeaxanthin, zinc) were tested on J774.2 cells. In particular, sub-confluent cells were pretreated (2 h) with AVS or individual components. Following LPS (1 µg/ml) treatment and overnight incubation, the medium was collected and nitrites and PGE₂ accumulation determined. Total RNA was also extracted to assess the expression of iNos and COX-2 mRNAs by Real Time RT-PCR. Data represent the average of 6 replicates from 3 different experiments. Statistical analysis were performed using one-way ANOVA plus Dunnett’s post hoc test.

Results : LPS induced the accumulation of ∼24 µmol/l nitrites and of ∼21 ng/ml PGE₂. Treatment with AVS, bromelain, zinc, salicin or anthocyanin effectively inhibited the accumulation of nitrites by 63-90% (p≤0.0001). Similarly AVS, bromelain or anthocyanin significantly inhibited PGE₂ accumulation by 38-99% (p≤0.05, p≤0.0001). The inhibition of nitrites and PGE₂ accumulation was paralleled by a significant inhibition of iNos (∼95%) and COX-2 (60-85%) expression, respectively. Components inactive when tested individually showed a synergistic effect on inhibition of nitrites and PGE₂ accumulation when mixed together (MixJ). This effect was paralleled by a significant inhibition of iNos expression (94%) while COX-2 inhibition (55%) failed to reach statistical significance.

Conclusions : The AVS formula was effective in inhibiting nitrites and PGE₂ accumulation in J774.2 cells. These activities appear to rely upon inhibition of iNos and COX-2 expression. Interestingly, components found to be inactive when tested individually were found to act synergistically when tested as a mix. Therefore, the AVS nutritional formula appears to be endowed with anti-inflammatory properties that may well prove useful to support anti-VEGF therapy.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.