HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Yang, Y. Articles by Awasthi, Y. C. Search for Related Content PubMed PubMed Citation Articles by Yang, Y. Articles by Awasthi, Y. C.

Protection of HLE B-3 Cells against Hydrogen Peroxide– and Naphthalene-Induced Lipid Peroxidation and Apoptosis by Transfection with hGSTA1 and hGSTA2

¹ From the Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, Texas; the ² Department of Ophthalmology and Visual Sciences, Washington University School of Medicine, St. Louis, Missouri; and the ³ Department of Chemistry and Biochemistry, University of Texas at Arlington, Arlington, Texas.

PURPOSE. To investigate the physiological role of two major -class glutathione S-transferases (GSTs), hGSTA1-1 and hGSTA2-2 in protection against oxidative stress and lipid peroxidation (LPO) in human lens epithelial (HLE B-3) cells.

METHODS. Total GSTs were purified from HLE B-3 cells by glutathione (GSH)-affinity chromatography and characterized by Western blot analysis, isoelectric focusing, and kinetic studies. The relative contributions of the -class GSTs and the Se-dependent glutathione peroxidase (GPx)-1 in GSH-dependent reduction of phospholipid hydroperoxide (PL-OOH) were quantitated through immunoprecipitation studies using separately the specific polyclonal antibodies against human -class GSTs and GPx-1. HLE B-3 cell membranes were prepared, peroxidized, and used to examine whether hGSTA1-1 and hGSTA2-2 catalyzes the reduction of membrane PL-OOH in situ using the microiodometric and spectrophotometric assays. The protective effects of the -class GSTs against H₂O₂- and naphthalene-induced LPO and apoptosis were examined by transfecting HLE B-3 cells with cDNAs of hGSTA1 and hGSTA2.

RESULTS. HLE B-3 cells expressed only the and class GSTs. The Michaelis-Menten constant (k_m) and turnover number (k_cat) of purified total GSTs toward phosphatidylcholine hydroperoxide (PC-OOH) were found to be 30 ± 4 µM and 1.95 ± 0.26 seconds, respectively. The -class GSTs accounted for approximately 65% of the total GPx activity of HLE B-3 cells toward PC-OOH. Our results demonstrate for the first time that hGSTA1-1 and hGSTA2-2 effectively catalyzed GSH-dependent reduction of membrane PL-OOH in situ in HLE B-3 cells. Transfection with hGSTA1 or hGSTA2 protected these cells from H₂O₂- and naphthalene-induced LPO and attenuated H₂O₂- and naphthalene-induced apoptosis through inhibiting caspase 3 activation.

CONCLUSIONS. These results demonstrate that the -class GSTs hGSTA1-1 and hGSTA2-2 play a major role as antioxidant enzymes and are the main determinants of the levels of LPO caused by oxidative stress in human lens epithelial cells.

This article has been cited by other articles:

				P. J. Boor, Y. Yang, and B. Gong Role of the Media in Vascular Injury: Atherosclerosis and Dissection Toxicol Pathol, January 1, 2006; 34(1): 33 - 38. [Abstract] [Full Text] [PDF]

				G. Chandrasekher and D. Sailaja Phosphatidylinositol 3-Kinase (PI-3K)/Akt but Not PI-3K/p70 S6 Kinase Signaling Mediates IGF-1-Promoted Lens Epithelial Cell Survival Invest. Ophthalmol. Vis. Sci., October 1, 2004; 45(10): 3577 - 3588. [Abstract] [Full Text] [PDF]

				M. Kantorow, J. R. Hawse, T. L. Cowell, S. Benhamed, G. O. Pizarro, V. N. Reddy, and J. F. Hejtmancik Methionine sulfoxide reductase A is important for lens cell viability and resistance to oxidative stress PNAS, June 29, 2004; 101(26): 9654 - 9659. [Abstract] [Full Text] [PDF]

				W. MA, D. LI, F. SUN, N. J. KLEIMAN, and A. SPECTOR The effect of stress withdrawal on gene expression and certain biochemical and cell biological properties of peroxide-conditioned cell lines FASEB J, March 1, 2004; 18(3): 480 - 488. [Abstract] [Full Text] [PDF]

				R. Sharma, Y. Yang, A. Sharma, S. Dwivedi, V. L. Popov, P. J. Boor, S. S. Singhal, S. Awasthi, and Y. C. Awasthi Mechanisms and Physiological Significance of the Transport of the Glutathione Conjugate of 4-Hydroxynonenal in Human Lens Epithelial Cells Invest. Ophthalmol. Vis. Sci., August 1, 2003; 44(8): 3438 - 3449. [Abstract] [Full Text] [PDF]

				S. Choudhary, S. Srivastava, T. Xiao, U. P. Andley, S. K. Srivastava, and N. H. Ansari Metabolism of Lipid Derived Aldehyde, 4-Hydroxynonenal in Human Lens Epithelial Cells and Rat Lens Invest. Ophthalmol. Vis. Sci., June 1, 2003; 44(6): 2675 - 2682. [Abstract] [Full Text] [PDF]

				A. Spector, D. Li, W. Ma, F. Sun, and P. Pavlidis Differential Amplification of Gene Expression in Lens Cell Lines Conditioned to Survive Peroxide Stress Invest. Ophthalmol. Vis. Sci., October 1, 2002; 43(10): 3251 - 3264. [Abstract] [Full Text] [PDF]