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Cameron K Postnikoff, Katherine S Held, Veena Viswanath, Kelly K Nichols; Closed eye tear neutrophils produce IL-17a in homeostatic conditions. Invest. Ophthalmol. Vis. Sci. 2018;59(9):147.
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Interleukin-17a (IL-17a) signaling has shown to be important in disease pathogenesis at the ocular surface, specifically in dry eye disease. While production of IL-17a has been mainly attributed to T cells, it has been demonstrated that neutrophils may also produce IL-17a in ocular surface fungal infection. Given that there are millions of leukocytes in closed eye tears, a pilot study was conducted to assess IL-17a production in these leukocytes for their potential relation to IL-17a regulation at the ocular surface.
Eight human participants were recruited and were trained to wash the ocular surface with phosphate buffered saline for at-home self-collection of tear leukocytes immediately following a full night of sleep (closed eye). Tear leukocytes were isolated and counted. Peripheral blood was collected by venipuncture into ethylenediaminetetraacetic acid vacutainer tubes. Blood leukocytes were separated by density gradient centrifugation using a combination of Polymorphprep (Axis-Shield; Oslo, Norway) and Histopaque (Sigma-Aldrich; St. Louis, MO). Unstimulated blood and tear leukocytes were incubated with a panel of fluorescently-labeled antibodies to distinguish T cells and neutrophils. A fixable viability stain was employed for exclusion of dead cells, as well as a dump channel to gate out B cell, monocyte, and natural killer cell contamination. Intracellular production of IL-17a was assessed using the transcription factor buffer set (BD Biosciences; San Jose, CA). Flow cytometry was performed using a BD LSR II.
On average, 0.4±0.2% of all unstimulated blood neutrophils were positive for IL-17a. In contrast, 10.0±9.0% of all unstimulated tear neutrophils were positive for IL-17a (p<0.01). While there were a small number of tear T cells that were positive for IL-17a (3.6±4.0%), the total number of IL-17a-producing tear neutrophils (1624±1513) was greater than total T cell counts measured in the collected tear samples (481±594) (p=0.05).
Intracellular IL-17a may be measured in unstimulated closed eye tear neutrophils. Closed eye tear neutrophils likely play a significant role in IL-17a production in the closed eye, which may be in part responsible for IL-17a dysregulation in dry eye disease.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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