GPR101 mediates the pro-resolving actions of RvD5n-3 DPA in arthritis and infections
Magdalena B. Flak, … , Francesco Palmas, Jesmond Dalli
Magdalena B. Flak, … , Francesco Palmas, Jesmond Dalli
Published January 2, 2020; First published December 3, 2019
Citation Information: J Clin Invest. 2020;130(1):359-373. https://doi.org/10.1172/JCI131609.
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Research Article Infectious disease Inflammation

GPR101 mediates the pro-resolving actions of RvD5n-3 DPA in arthritis and infections

Abstract

N-3 docosapentaenoic acid–derived resolvin D5 (RvD5n-3 DPA) is diurnally regulated in peripheral blood and exerts tissue-protective actions during inflammatory arthritis. Here, using an orphan GPCR screening approach coupled with functional readouts, we investigated the receptor(s) involved in mediating the leukocyte-directed actions of RvD5n-3 DPA and identified GPR101 as the top candidate. RvD5n-3 DPA bound to GPR101 with high selectivity and stereospecificity, as demonstrated by a calculated KD of approximately 6.9 nM. In macrophages, GPR101 knockdown limited the ability of RvD5n-3 DPA to upregulate cyclic adenosine monophosphate, phagocytosis of bacteria, and efferocytosis. Inhibition of this receptor in mouse and human leukocytes abrogated the pro-resolving actions of RvD5n-3 DPA, including the regulation of bacterial phagocytosis in neutrophils. Knockdown of the receptor in vivo reversed the protective actions of RvD5n-3 DPA in limiting joint and gut inflammation during inflammatory arthritis. Administration of RvD5n-3 DPA during E. coli–initiated inflammation regulated neutrophil trafficking to the site of inflammation, increased bacterial phagocytosis by neutrophils and macrophages, and accelerated the resolution of infectious inflammation. These in vivo protective actions of RvD5n-3 DPA were limited when Gpr101 was knocked down. Together, our findings demonstrate a fundamental role for GPR101 in mediating the leukocyte-directed actions of RvD5n-3 DPA.

Authors

Magdalena B. Flak, Duco S. Koenis, Agua Sobrino, James Smith, Kimberly Pistorius, Francesco Palmas, Jesmond Dalli

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Figure 2

Activation of GPR101 by RvD5n-3 DPA.

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Activation of GPR101 by RvD5n-3 DPA. (A) RvD5n-3 DPA was incubated at th...
(A) RvD5n-3 DPA was incubated at the indicated concentrations with CHO cells expressing human GPR101 (circles), GPR84 (squares), or GPR12 (triangles) coupled with the β-arrestin reporter system, and receptor activation was measured as an increase in luminescence signal. Results represent the mean ± SEM. n = 5–7 independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 versus the respective vehicle control group; 2-way ANOVA with Tukey’s post hoc multiple comparisons test. (B) CHO cells overexpressing GPR101 were incubated with either isotype control or anti-GPR101 antibody (30 minutes at room temperature) and then with 1 nM RvD5n-3 DPA, and impedance was measured over a 20-minute period using the xCELLigence DP system. Results are representative of 3 distinct experiments. (C) CHO cells expressing GPR101 coupled with the β-arrestin reporter system were incubated with the indicated concentrations of RvD5n-3 DPA, RvD1n-3 DPA, PD1n-3 DPA, or vehicle (PBS containing 0.01% ethanol), and receptor activation was measured as an increase in luminescence signal. Results represent the mean ± SEM. n = 5–7 independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 versus the vehicle control group; 2-way ANOVA with Tukey’s post hoc multiple comparisons test. (D) RvD5n-3 DPA, RvD1n-3 DPA, and PD1n-3 DPA (10 nM) were incubated with GPR101-expressing CHO cells, and impedance was measured over a 30-minute period using the xCELLigence DP system. Results are representative of 3 distinct experiments.