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Shuang Zhu, Xiaobing Hu, Yonju Ha, Hua Liu, Wenbo Zhang; ER stress induces CXCL10 and CCL2 expression in photoreceptors via common and distinct mechanisms. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):2392.
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© ARVO (1962-2015); The Authors (2016-present)
Photoreceptors are recently found as a key player in retinal diseases including diabetic retinopathy. However, it is largely unknown how photoreceptors interact with other cells during diseases. CCL2 and CXCL10 are two chemokines that play an important role in vascular inflammation by inducing leukocyte recruitment and activation. This study is to test the hypothesis that photoreceptors can produce CCL2 and CXCL10 under stress and to eliminate potential mechanisms of stress-induced CCL2 and CXCL10 production.
Studies were performed using RGC-5, a photoreceptor cell line closely related to photoreceptor cell line 661W. Cells were treated with thapsigargin (TG, 0.1 uM), an inducer of endoplasmic reticulum (ER) stress, and levels of CXCL10, CCL2, ER stress markers, and MAPKs were measured by qPCR, ELISA, and/or Western blot. ER stress pathways, MAPKs and NADPH oxidase were blocked by siRNAs or inhibitors.
At 24 hour after TG treatment, CCL2 and CXCL10 proteins were increased by 2.4 fold and 3.3 fold, respectively. Both CXCL10 and CCL2 mRNAs were significantly elevated at 1-24 hour after treatment. However, CCL2 mRNA rapidly declined after it reached peak increase (44.1 fold) at 3 hour after TG treatment whereas CXCL10 mRNA continuously increased from 9.8 fold to 17.1 fold in the period of 3-24 hour after treatment. TG-induced CXCL10 and CCL2 expression was significantly blocked by phenylbutyric acid, an ER stress blocker. ER stress contains three pathways: PERK, ATF6 and IRE1a. While all of them can induce CHOP expression, transcription factor Xbp1s is only induced by IRE1a. Knockdown of PERK reduced TG-induced CXCL10 and CCL2 expression by 45.2% and 80.6%, respectively. Although knockdown of ATF6 and CHOP significantly reduced TG-induced CXCL10 expression and knockdown of Xbp1s increased CXCL10 expression, none of them affected TG-induced CCL2 expression. TG-induced CXCL10 and CCL2 were also robustly blocked by p38MAPK inhibitor (SB202190) and NADPH oxidase inhibitor (DPI) but not by ERK and JNK inhibitors.
These studies suggest that photoreceptors may be involved in retinal inflammatory reactions during diseases by expressing chemokines CXCL10 and CCL2. Multiple pathways during ER stress were involved in regulating CXCL10 expression whereas CCL2 was mainly modulated by PERK. Additionally, p38MAPK and NADPH oxidase were involved in CXCL10 and CCL2 expression.
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