The protective effect of functional connexin43 channels on a human epithelial cell line exposed to oxidative stress.

Abstract:

PURPOSE:To determine the role of connexin43 (Cx43) and gap junctional intercellular communication (GJIC) in the response of the human retinal pigment epithelial cell line ARPE-19 to oxidative stress. METHODS:ARPE-19 cells were treated with the chemical oxidant tert-butyl hydroperoxide (t-BOOH), and cell viability was assessed by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. GJIC was evaluated by scrape loading/dye transfer and microinjection assays, and Cx43 expression was detected by Western blot and immunofluorescent staining combined with confocal microscopy analysis. Retroviral infection of ARPE-19 cells with shRNA vectors targeting Cx43 or vectors encoding Cx43, Cx26, and a disease-linked dominant negative Cx43 mutant (G21R) were used, and the effect on cell viability was assessed. RESULTS:t-BOOH-induced ARPE-19 cell death was correlated with reductions in GJIC and in the total level of Cx43 protein expression. Overexpression of Cx26 and Cx43 increased the viability of oxidant-treated ARPE-19 cells. Conversely, shRNA knockdown of Cx43, expression of a disease-linked dominant negative Cx43 mutant, and blocking GJIC with 18beta-glycyrrhetinic acid and flufenamic acid all increased t-BOOH-induced ARPE-19 cell death. CONCLUSIONS:Cx43-mediated protection of ARPE-19 cells from oxidative stress-induced death is dependent on functional Cx43 channels.

authors

Hutnik CM,Pocrnich CE,Liu H,Laird DW,Shao Q

doi

10.1167/iovs.07-0717

subject

Has Abstract

pub_date

2008-02-01 00:00:00

pages

800-6

issue

2

eissn

0146-0404

issn

1552-5783

pii

49/2/800

journal_volume

49

pub_type

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