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Hypoxia induces an inflammatory response in ARPE ‐19 cells
Author(s) -
Arjamaa O.,
Aaltonen V.,
Piippo N.,
Kaarniranta K.,
Kauppinen A.
Publication year - 2015
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2015.0426
Subject(s) - hypoxia (environmental) , secretion , intracellular , cell culture , interleukin 8 , inflammation , chemistry , microbiology and biotechnology , immunology , biology , pharmacology , andrology , medicine , biochemistry , oxygen , organic chemistry , genetics
Purpose To characterize further the link between hypoxia and inflammation in retinal diseases, we studied here the effects of hypoxic exposure on the secretion of inflammatory cytokines from ARPE ‐19 cell culture, pretreated with lipoplysaccharide ( LPS , to induce inflammation). Methods ARPE cell culture was exposed to 24 h hypoxia, 24 h hypoxia followed by 24 h reoxygenation, and 24 h or 48 h normoxia with or without LPS . Experiments were performed using culture medium DMEM /F‐12 supplemented with penicillin, streptomycin and L‐glutamin without or serum and with or without LPS . Each treatment was repeated 6 times. Hypoxia (37 C, 1% O2, 5% CO 2, 90% moisture) was induced in Ruskinn Invivo2 workstation, and normoxia and reoxygenation (37C, air, 5% CO 2, 90% moisture) in a standard cell culture incubator. The culture media and cell lysates were collected under hypoxic or normoxic atmosphere, centrifuged and snap frozen for storage and further analyses. VEGF and cytokines were measured with ELISA and intracellular proteins (autophagy markers p62, LC 3 and oxidative stress marker Nrf2) with immunoblotting. Results As expected, hypoxic conditions increased significantly the secretion of VEGF from ARPE ‐19 cell culture as compared to the secretion in normoxic conditions. Also, the secretion of IL ‐6 and IL ‐8 showed a significant increase in hypoxia when measured at 24 h. The intracellular protein levels were changed: p62 increased while LC 3 and Nrf2 decreased in hypoxia (at 6 h). Conclusions An acute exposure to hypoxia induced an inflammatory response in ARPE ‐19 cell culture as characterized with an increased IL ‐6 and IL ‐8 secretion from cell culture. Intracellularly, the autophagic response decreased (seen with the increase of p62 and with the decrease of LC 3) and the oxidative stress increased (seen with the decrease of Nrf2).