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. 2017 Sep 8;7:11009. doi: 10.1038/s41598-017-11188-y

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Distinct SPM profiles in IL-1β stimulated healthy and diseased tendon stromal cells. Tendon stromal cells (60,000 cells per well) were derived from healthy hamstring (n = 8 donors) or diseased supraspinatus tendons (n = 8 donors). Cells were cultured in DMEM F12 phenol red free medium containing 1% heat inactivated human serum to 80% confluence and incubated with IL-1β for 24 h. Media and cells were harvested and placed in ice-cold methanol containing deuterium labeled internal standards. LM were then extracted and profiled. (A) 2-dimensional score plot and (B) corresponding 2-dimensional loading plot of plasma LM-SPM from human tendon derived-stromal cell incubations isolated from healthy volunteers or patients with tendinopathy after stimulation with IL-1β (10ngml^—1) for 24 h. Grey ellipse in the score plot denotes 95% confidence regions. (C) Concentrations for mediators found to be differentially regulated between healthy (grey bars) and diseased (black bars) tendon stromal cell incubations. Results are shown as means and SEM and representative of n = 8 donors per group. (D) mRNA expression of lipid mediator biosynthetic enzymes determines using real time qPCR. Gene expression is normalized to β-actin, bars show median values. (E) Representative immunofluorescence images showing staining for ALOX15 (green), PTGS2 (red) and nuclei (cyan) in IL-1β-stimulated healthy and diseased tendon cells. Scale bar, 20μm. Results are representative of n = 3 donors.