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Jing Wu, Coralia Luna, Guorong Li, David L. Epstein, Pedro Gonzalez; Alterations In Arachidonic Acid Metabolism Induced By Mechanical And Oxidative Stress In Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3256.
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We have previously reported that cyclic mechanical stress (CMS) lead to mobilization of arachidonic acid (AA) from the plasma membrane in trabecular meshwork (TM) cells. These results suggested that CMS might induce the production of bioactive lipids capable of influencing the physiology of the outflow pathway. Here we used a targeted lipidomics approach to evaluate the effects of CMS and oxidative stress on the production of specific AA derivatives by porcine TM cells.
Primary cultures of porcine TM cells at passage 3 were subjected to CMS (20% stretching 1 hertz frequency) for 1 and 3 hours using a Flexcell system. For chronic oxidative stress cells were incubated at oxygen concentrations of 40% (stressed) or 5% (control) for 8 days. Targeted lipidomic analysis of specific AA derivatives (PGE2, PGF2a, 14,15-DHET, 15-HETE, 8-iso-PGE2) was conducted by HPLC-tandem mass spectrometry in media samples from experimental and control cultures.
The more abundant AA derivatives detected in medium from control TM cells 1 hour after media change were PGE2 (0.099 ng/mL); PGF2a (0.019 ng/mL); 14,15-DHET (0.017 ng/mL); and 15-HETE (0.014 ng/mL). CMS lead to a marked increase in of PGE2 (5.6±0.27 fold, p<0.01) and 8-iso-PGE2 (20±3 fold, p<0.01) and a smaller increase (58±22%, p<0.05) in PGF2a. Incubation of confluent cultures of porcine TM cells 40% oxygen for 8 days resulted in a significant decrease in concentration of PGE2 (35±0.7%, p<0.01), PGF2a (34±3.9%, p=0.001), 8-iso-PGE2 (41±2.8%, p<0.05) and 14,15-DHET (49±1.5%, p<0.01), and a significant increase (358±20%, p<0.001) of 15(s)-HETE compared to control cultures incubated at 5% oxygen.
The observed increase in PGE2 and other AA derivatives induced by CMS in TM cells could potentially constitute a homeostatic response aimed at increasing aqueous humor outflow facility in response to increases in IOP. Chronic oxidative stress might in turn contribute to the pathogenesis of the TM in glaucoma by shifting the metabolism of AA from the synthesis of IOP-lowering molecules such as PGE2 to the production of non-enzymatically oxidized AA derivatives.
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