TPA-induced inhibition of gap junctional intercellular communication is not mediated through free radicals.
The present investigation was designed to determine whether the inhibition of gap junction-mediated intercellular communication (GJIC) induced by TPA (12-O-tetradecanoylphorbol-13-acetate) in rat liver epithelial (WB-F344) cells in vitro is mediated through free radical production. As assessed by fluorescence redistribution after photobleaching (FRAP) analysis, GJIC was significantly inhibited in cells treated for 1 hr with either 10 ng/ml TPA or 500 microM hydrogen peroxide (H2O2). Addition of 1000 U/ml catalase or 25 microM N',N'-diphenyl-p-phenylenediamine (DPPD) to TPA-treated cells did not alleviate the TPA-induced inhibition of GJIC. However, the concurrent addition of 1000 U/ml catalase to the culture medium prevented the H2O2 inhibition of GJIC. 2'-7'-dichlorofluorescein-mediated fluorescence, a measure of free radical production utilizing the Meridian ACAS 470 interactive laser cytometer, was not significantly increased in WB-F344 cells treated with 10 and 100 ng/ml TPA when compared to control cells. However, polymorphonuclear leukocytes (PMNs) treated for 10 min with 100 ng/ml TPA showed a substantial oxidative burst, as did WB-F344 cells treated for 1 hr with 500 microM H2O2. The concurrent addition of 1000 U/ml catalase to the culture medium attenuated H2O2-mediated free radical production in both PMNs and WB-F344 cells. Data from this study do not support a role for free radicals in the TPA-induced inhibition of GJIC in WB-F344 cells.
Hasler
C M
CM
Department of Food Science and Human Nutrition, Michigan State University, East Lansing 48824.
Frick
M A
MA
Bennink
M R
MR
Trosko
J E
JE
D000818:Animals; D002417:Cattle; D002450:Cell Communication / Q000187:drug effects*; D002478:Cells, Cultured; D004396:Coloring Agents; D015536:Down-Regulation / Q000187:drug effects; D004847:Epithelial Cells; D005453:Fluorescence; D005609:Free Radicals; D007365:Intercellular Junctions / Q000187:drug effects* / Q000502:physiology; D008099:Liver / Q000166:cytology; D008297:Male; D008722:Methods; D010777:Photochemistry; D051381:Rats; D011919:Rats, Inbred Strains; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D004396:Coloring Agents; D005609:Free Radicals; D013755:Tetradecanoylphorbol Acetate