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Sumin Li, Yin Wang, Marjorie F. Lou; Thioredoxin 1 stimulates proliferation of human lens epithelial cells by activating NF-B pathway. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1067.
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Thioredoxin 1(Trx1) is an important intracellular protein that modulates redox balance through catalyzing dithiol/disulfide exchange reactions. Our previous studies showed that exogenous human Trx1 induced reactive oxygen species generation and stimulated proliferation in human lens epithelial (HLE) B3 cells through activations of MAPK and AKT (survival signaling factor) pathways. The current study is to investigate if NF-kB, a known transcription factor in controlling cell proliferation, may involve in Trx1-induced cell growth.
HLE B3 cells were cultured with 20 μM Trx1 for multiple time points (5-240 min) and the cell lysates were separated into cytosolic and nuclear fractions. Both fractions were examined for the activation or phosphorylation of ERK1/2, AKT and IKBα (NF-kB inhibitor), and the expression of NF-ΚB using Western blot analysis. Electrophoretic mobility shift assay (EMSA) was employed to determine NF-ΚB-DNA binding ability. Inhibitor to MEK1/2 (U0126) or to PI3K/AKT (LY294002) was each added to the culture 30 min prior to Trx1 treatment to test the possible role of ERK1/2 or AKT pathway in Trx1-induced NF-ΚB signaling.
Exogenous Trx1 transiently induced P-ERK1/2, P-AKT and nuclear translocated P-ERK1/2 with a peak stimulation at 10 min post-treatment. However, Trx1 showed a delayed effect on IKBα phosphorylation at 30-60 min, followed by nuclear translocation of NF-ΚB that lasted from 60-240 min. EMSA confirmed that NF-ΚB-DNA binding was correspondingly enhanced in response Trx1 stimulation. Neither LY294002 nor U0126 affected NF-ΚB translocation and its DNA-binding ability induced by Trx1.
Trx1 stimulated cell proliferation may be in part mediated through NF-ΚB activation that is independent of ERK and PI3K pathways.
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