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Free fatty acid receptor 4 activation protects against choroidal neovascularization in mice

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To examine whether free fatty acid receptor 4 (FFAR4) activation can protect against choroidal neovascularization (CNV), which is a common cause of blindness, and to elucidate the mechanism underlying the inhibition, we used the mouse model of laser-induced CNV to mimic angiogenic aspects of age-related macular degeneration (AMD). Laser-induced CNV was compared between groups treated with an FFAR4 agonist or vehicle, and between FFAR4 wild-type (Ffar4+/+) and knock out (Ffar4−/−) mice on a C57BL/6J/6N background. The ex vivo choroid-sprouting assay, including primary retinal pigment epithelium (RPE) and choroid, without retina was used to investigate whether FFAR4 affects choroidal angiogenesis. Western blotting for pNF-ĸB/NF-ĸB and qRT-PCR for Il-6, Il-1β, Tnf-α, Vegf, and Nf-ĸb were used to examine the influence of FFAR4 on inflammation, known to influence CNV. RPE isolated from Ffar4+/+ and Ffar4−/− mice were used to assess RPE contribution to inflammation. The FFAR4 agonist suppressed laser-induced CNV in C57BL/6J mice, and CNV increased in Ffar4−/− compared to Ffar4+/+ mice. We showed that the FFAR4 agonist acted through the FFAR4 receptor. The FFAR4 agonist suppressed mRNA expression of inflammation markers (Il-6, Il-1β) via the NF-ĸB pathway in the retina, choroid, RPE complex. The FFAR4 agonist suppressed neovascularization in the choroid-sprouting ex vivo assay and FFAR4 deficiency exacerbated sprouting. Inflammation markers were increased in primary RPE cells of Ffar4−/− mice compared with Ffar4+/+ RPE. In this mouse model, the FFAR4 agonist suppressed CNV, suggesting FFAR4 to be a new molecular target to reduce pathological angiogenesis in CNV.

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The work was supported by Grants Manpei Suzuki Diabetic Foundation (YT), The German Research Foundation (DFG; to BC [CA1940/1-1]), Boston Children's Hospital OFD/BTREC/CTREC Faculty Career Development Grant, Boston Children's Hospital Ophthalmology Foundation, Little Giraffe Foundation, BCH Manton Center Fellowship and Blind Children’s Center (ZF), NIH/NEI (Grant Nos. R01EY030140, R01EY029238), Bright Focus Foundation, Boston Children's Hospital Ophthalmology Foundation (YS), The Wallenberg Clinical Scholars (AH), The Vascular Biology Program and the Boston Children’s Hospital Surgery Foundation (MP), and NIH R24EY024868, EY017017, R01EY01717-13S1, EY030904-01, BCH IDDRC (Grant No. 1U54HD090255), Massachusetts Lions Eye Foundation (LEHS).

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YT, BC, ZF, YS, MP, AH, ST, LEHS. designed all experiments; YT, BC, ZF, CHL, SH, SC, WB, performed the experiments; YT analyzed the data; YT, BC and LEHS wrote the manuscript.

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Correspondence to Lois E. H. Smith.

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The authors declare that there is no duality of interest associated with this manuscript. Saswata Talukdar is an employee and stockholder of Merck &Co.

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All animal studies adhered to the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Vision Research and were approved by Institutional Animal Care and Use Committee at Boston Children’s Hospital (ARCH Protocol Number 19-04-3913R).

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Tomita, Y., Cakir, B., Liu, CH. et al. Free fatty acid receptor 4 activation protects against choroidal neovascularization in mice. Angiogenesis 23, 385–394 (2020).

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