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. 2020 Sep 4;9(9):828.
doi: 10.3390/antiox9090828.

A Higher Proportion of Eicosapentaenoic Acid (EPA) When Combined with Docosahexaenoic Acid (DHA) in Omega-3 Dietary Supplements Provides Higher Antioxidant Effects in Human Retinal Cells

Manuel Saenz de Viteri  1   2   3   4 , María Hernandez  2   3 , Valentina Bilbao-Malavé  1   2   3 , Patricia Fernandez-Robredo  2   3   4 , Jorge González-Zamora  1   2   3 , Laura Garcia-Garcia  2 , Nahia Ispizua  2 , Sergio Recalde  2   3   4 , Alfredo Garcia-Layana  1   2   3   4
Affiliations

A Higher Proportion of Eicosapentaenoic Acid (EPA) When Combined with Docosahexaenoic Acid (DHA) in Omega-3 Dietary Supplements Provides Higher Antioxidant Effects in Human Retinal Cells

Manuel Saenz de Viteri et al. Antioxidants (Basel). .

Abstract

Retinal pigment epithelium (RPE) is a key regulator of retinal function and is directly related to the transport, delivery, and metabolism of long-chain n-3 polyunsaturated fatty acids (n3-PUFA), in the retina. Due to their functions and location, RPE cells are constantly exposed to oxidative stress. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have shown to have antioxidant effects by different mechanisms. For this reason, we designed an in vitro study to compare 10 formulations of DHA and EPA supplements from different origins and combined in different proportions, evaluating their effect on cell viability, cell proliferation, reactive oxygen species production, and cell migration using ARPE-19 cells. Furthermore, we assessed their ability to rescue RPE cells from the oxidative conditions seen in diabetic retinopathy. Our results showed that the different formulations of n3-PUFAs have a beneficial effect on cell viability and proliferation and are able to restore oxidative induced RPE damage. We observed that the n3-PUFA provided different results alone or combined in the same supplement. When combined, the best results were obtained in formulations that included a higher proportion of EPA than DHA. Moreover, n3-PUFA in the form of ethyl-esters had a worse performance when compared with triglycerides or phospholipid based formulations.

Keywords: diabetic retinopathy; docosahexaenoic acid (DHA); eicosapentaenoic acid (EPA); oxidative stress; retinal pigment epithelium.

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Conflict of interest statement

AGL is a consultant for Bayer, Novartis, Allergan, Thea, and Roche. The rest of the authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Figures

Figure 1
Figure 1
Effect of omega-3 supplements on epithelial tight junctions. ZO-1 (red) immunofluorescence were not affected by any of the omega-3 treatments. Nuclei are stained with TOPRO-3. Scale bar 20 µm. (A): Control group; (B): H2O2 treatment group; (C): DHA (Docosahexaenoic acid) group; (D): EPA (Eicosapentaenoic acid) group; (E): EPA/DHA 40/20; TG (Triglycerides) group; (F): EPA/DHA 20/40 TG group; (G): EPA/DHA 40/20 EE (Ethylesters) group; (H): DHA 97/3V (Vegetable) group; (I): DHA 95/5V group; (J): DHA 97/3M (Marine) group; (K): DHA 97/3 VM 1.5 group; (L): DHA 95/5 VM 2.5 group.
Figure 2
Figure 2
Graphs showing relative cell viability (A–C) and proliferation (D–F) compared to a control group under normal conditions of groups 1, 2, and 3 of eicosapentaenoic acid/docosahexaenoic acid (DHA/EPA) formulations, respectively (D–F). For comparisons, one-way ANOVA with the Bonferroni post-hoc test were used. Data are expressed as mean ± SEM. * p < 0.05. EPA: Eicosapentaenoic acid; DHA: Docosahexaenoic acid; TG: Triglycerides; EE: Ethylesters; PL: Phospholipids. V: Vegetable. M: Marine.
Figure 3
Figure 3
Graphs showing relative cell viability (A–C) and proliferation (D–F) results for ARPE-19 cells treated with different DHA and EPA treatments groups under oxidative stress conditions (H2O2 For comparisons, one-way ANOVA with the Bonferroni post-hoc test were used. Data are expressed as mean ± SEM. * p < 0.05 and ** p < 0.01. EPA: Eicosapentaenoic acid; DHA: Docosahexaenoic acid; TG: Triglycerides; EE: Ethylesters; PL: Phospholipids. V: Vegetable. M: Marine.
Figure 4
Figure 4
Graphs showing relative Dichloride fluoresceine (DCF) detection results for ARPE-19 cells in groups 1, 2, and 3 of DHA/EPA formulations, respectively (A–C). For comparisons, one-way ANOVA with the Bonferroni post-hoc test were used. Data are expressed as mean ± SEM. ** p < 0.01 *** p < 0.001. EPA: Eicosapentaenoic acid; DHA: Docosahexaenoic acid; TG: Triglycerides; EE: Ethylesters; PL: Phospholipids. V: Vegetable. M: Marine.
Figure 5
Figure 5
Graphs showing relative caspase-3 expression (positive granules per nucleus) (A–C) results for ARPE-19 cells treated with different groups 1, 2, and 3 of DHA/EPA formulations, respectively. For comparisons, one-way ANOVA with the Bonferroni post-hoc test were used. (D) Scale bar 20 µm Data are expressed as mean ± SEM. EPA: Eicosapentaenoic acid; DHA: Docosahexaenoic acid; TG: Triglycerides; EE: Ethylesters; PL: Phospholipids. V: Vegetable. M: Marine.
Figure 6
Figure 6
Graphs showing relative migration ratio (A–C) results for ARPE-19 cells treated with different groups 1, 2, and 3 of DHA/EPA formulations, respectively. For comparisons, one-way ANOVA with the Bonferroni post-hoc test were used. Data are expressed as mean ± SEM. ** p < 0.01. EPA: Eicosapentaenoic acid; DHA: Docosahexaenoic acid; TG: Triglycerides; EE: Ethylesters; PL: Phospholipids. V: Vegetable. M: Marine.
Figure 7
Figure 7
Graphs showing relative VEGF/PEDF ratio (A–C) results for ARPE-19 cells treated with different groups 1, 2, and 3 of DHA/EPA formulations, respectively. For comparisons, one-way ANOVA with the Bonferroni post-hoc test were used. Data are expressed as mean ± SEM. EPA: Eicosapentaenoic acid; DHA: Docosahexaenoic acid; TG: Triglycerides; EE: Ethylesters; PL: Phospholipids. V: Vegetable. M: Marine.
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