12(S)-HETE mediates diabetes-induced endothelial dysfunction by activating intracellular endothelial cell TRPV1
Mandy Otto, … , Eric R. Gross, Nana-Maria Wagner
Mandy Otto, … , Eric R. Gross, Nana-Maria Wagner
Published June 25, 2020
Citation Information: J Clin Invest. 2020;130(9):4999-5010. https://doi.org/10.1172/JCI136621.
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12(S)-HETE mediates diabetes-induced endothelial dysfunction by activating intracellular endothelial cell TRPV1

Abstract

Patients with diabetes develop endothelial dysfunction shortly after diabetes onset that progresses to vascular disease underlying the majority of diabetes-associated comorbidities. Increased lipid peroxidation, mitochondrial calcium overload, and mitochondrial dysfunction are characteristics of dysfunctional endothelial cells in diabetic patients. We here identified that targeting the lipid peroxidation product 12(S)-hydroxyeicosatetraenoic acid–induced [12(S)-HETE–induced] activation of the intracellularly located cation channel transient receptor potential vanilloid 1 (TRPV1) in endothelial cells is a means to causally control early-stage vascular disease in type I diabetic mice. Mice with an inducible, endothelium-specific 12/15-lipoxygenase (12/15Lo) knockout were protected similarly to TRPV1-knockout mice from type 1 diabetes–induced endothelial dysfunction and impaired vascular regeneration following arterial injury. Both 12(S)-HETE in concentrations found in diabetic patients and TRPV1 agonists triggered mitochondrial calcium influx and mitochondrial dysfunction in endothelial cells, and 12(S)-HETE effects were absent in endothelial cells from TRPV1-knockout mice. As a therapeutic consequence, we found that a peptide targeting 12(S)-HETE–induced TRPV1 interaction at the TRPV1 TRP box ameliorated diabetes-induced endothelial dysfunction and augmented vascular regeneration in diabetic mice. Our findings suggest that pharmacological targeting of increased endothelial lipid peroxidation can attenuate diabetes-induced comorbidities related to vascular disease.

Authors

Mandy Otto, Clarissa Bucher, Wantao Liu, Melanie Müller, Tobias Schmidt, Marina Kardell, Marvin Noel Driessen, Jan Rossaint, Eric R. Gross, Nana-Maria Wagner

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Figure 5

Abrogation of the 12(S)-HpETE/TRPV1 interaction protects against diabetes-induced endothelial dysfunction.

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Abrogation of the 12(S)-HpETE/TRPV1 interaction protects against diabete...
(A) Schematic of TRPV1 corresponding to amino acids 701–711 within the TRP box at the C-terminus of human TRPV1 linked to a TAT linker protein for intracellular entry. (B) Patch clamp experiments show that 700 nM 12(S)-HpETE evokes currents in endothelial cells loaded with 1 μM TAT linker, but not 1 μM V1-cal. Representative patch results from n = 10 independent experiments. (C) 12(S)-HpETE–induced mitochondrial calcium changes detected by flow cytometry [time point of addition of 12(S)-HpETE is indicated by the arrow] in human endothelial cells preincubated with either 1 μM V1-cal or 1 μM of a scrambled version of V1-cal (V1-scr) or TAT linker protein serving as controls. ***P < 0.001 vs. corresponding time point in V1-cal or EtOH, #P < 0.05, ##P < 0.01, ###P < 0.001 vs. TAT or V1-scr, 2-way ANOVA/Bonferroni, n = 5 independent experiments. (D) Assessment of mitochondrial function to evaluate effects of 1 μM V1-cal versus 1 μM TAT linker only (top 2 panels) or 1 μM V1-scr (bottom panel) on mitochondrial OCR induced by 1 μM 12(S)-HpETE. After 3 measurements, oligomycin was added before FCCP used to induce maximum respiration. *P < 0.05, **P < 0.01, 2-way ANOVA/Bonferroni. Variants were analyzed in triplicate on Seahorse miniplates, and n = 5 independent experiments were performed. All graphs show mean ± SEM. (E) Cch-induced vasorelaxation shows that repetitive intravenous application on 4 consecutive days of the V1-cal peptide (1 mg/kg/d), but not of the V1-scr peptide or the TAT linker protein only, to diabetic WT mice reduced endothelial dysfunction induced by diabetes (STZ). Sodium citrate (NaCi) was used as vehicle control. ***P < 0.001 vs. as indicated, 2-way ANOVA/Bonferroni. Means were generated for each individual animal by analysis of 2 arteries; n = 5 animals per group. Graph shows mean ± SEM.