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2014 ; 55
(12
): 7961-9
Nephropedia Template TP
Tibrewal S
; Ivanir Y
; Sarkar J
; Nayeb-Hashemi N
; Bouchard CS
; Kim E
; Jain S
Invest Ophthalmol Vis Sci
2014[Nov]; 55
(12
): 7961-9
PMID25406284
show ga
PURPOSE: To determine if hyperosmolar stress can stimulate human neutrophils to
form neutrophil extracellular traps (NETs) and to investigate potential
strategies to reduce formation of NETs (NETosis) in a hyperosmolar environment.
METHODS: Neutrophils were isolated from peripheral venous blood of healthy
subjects and incubated in iso-osmolar (280 mOsM) or hyperosmolar (420 mOsM) media
for 4 hours. Neutrophil extracellular traps were quantified using a PicoGreen dye
assay to measure extracellular DNA. Two known inhibitors of NETosis,
staurosporine and anti-?2 integrin blocking antibody, and two proresolution
formyl peptide receptor 2 (FPR2) agonists, annexin/lipocortin-1 mimetic peptide
and 15-epi-lipoxin A4, were evaluated as possible strategies to reduce
hyperosmolarity-induced NETosis. RESULTS: The amount of NETs induced by
hyperosmolar medium (420 mOsM) increased linearly over time to 3.2 ± 0.3 times
that induced by iso-osmolar medium at 4 hours (P < 0.05). NETosis increased
exponentially with increasing osmolarity and was independent of the stimulus used
to increase osmolarity. Upon neutrophil exposure to hyperosmolar stress,
restoration of iso-osmolar conditions decreased NET formation by 52.7% ± 5% (P <
0.05) but did not completely abrogate it. Among the strategies tested to reduce
NETosis in a hyperosmolar environment, annexin-1 peptide was the most
efficacious. CONCLUSIONS: Hyperosmolarity induces formation of NETs by
neutrophils. This NETosis mechanism may explain the presence of excessive NETs on
the ocular surface of patients with dry eye disease. Because they reduce
hyperosmolarity-induced NETosis, FPR2 agonists may have therapeutic potential in
these patients.