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MF Lou, K Krysan; The Presence of a Redox Signaling System In The Lens . Invest. Ophthalmol. Vis. Sci. 2002;43(13):848.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: A physiological role of reactive oxygen species (ROS), the redox signaling, was identified in other tissues, which indicates that exogenous ROS at low levels could mimic certain growth factors to regulate and activate the mitogenic and stress-associated signal transduction pathways in the cells and lead to gene expressions for various cellular functions. This study is to verify if the redox signaling system is present in the lens. Methods: Confluent human lens epithelial cells (B3) (1.6 millions) were waned of serum by first culturing in 2% fetal bovine serum overnight and then in serum-free medium for 30 min before subjected to a bolus of H2O2 (0.1 mM) for 0, 5, 15, 30 and 60 min in the presence and absence of genistein, a protein tyrosine kinase inhibitor. Untreated cells were used as controls. The cells were lysed immediately on plates with modified RIPA buffer (50 mM Tris-HCl, pH 7.4; 1% NP-40; 150 mM NaCl; 1 mM EDTA; 1 mM PMSF; 1 mM NaF; 1mM Na3VO4; 0.25% Na-deoxycholate; 1 µg/ml each of aprotinin, leupeptin and pepstatin). The cell lysates were examined for the activation of mitogenic activated protein kinases (MAPK) Raf-1-MEK-ERK cascade, the stress-associated MAPK (p38 and JNK) and the survival factor, Akt, by using Western blot analysis with respective phospho-specific antibobies. Results: We observed a transient stimulation of the Raf-MEK-ERK cascade as well as JNK and p38. The activation began within 5 min after H2O2 exposure and peaked by 10-15 mins before gradually subsiding. By 60 min, phosphorylation of some kinases (MEK and p38) returned to basal levels. Akt had a delayed response, which only began to activate at 30 min and was deactivated by 60 min. These time-dependent patterns of transient stimulation correlated with the dissipation pattern of H2O2 in the medium (detoxification by the cells). Genistein eliminated the effect of H2O2 in Raf, p38 and Akt completely and diminished the activation of MEK, ERK and JNK substantially. Conclusion: The redox signaling system is present in the lens since the HLE cells showed strong response to H2O2 stimulation, which is mediated through protein tyrosine kinase, similar to that of bFGF or EGF growth factors.
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