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Regulation of thioltransferase expression in human lens epithelial cells.
Invest Ophthalmol Vis Sci. 2001 Apr; 42(5):1002-8.IO

Abstract

PURPOSE

To study how the expression of thioltransferase (TTase), a critical thiol repair and dethiolating enzyme, is regulated in human lens epithelial cells under oxidative stress. Also to examine whether depleting the primary cellular antioxidant glutathione (GSH) in these cells has any influence on TTase expression under the same conditions.

METHODS

Human lens epithelial cells (B3) were grown to confluence (1.6 million) and gradually weaned from serum in the medium before exposing to 0.1 mM H2O2 for 2 hours. Cells were removed at the time intervals of 0, 5, 10, 15, 30, 60, and 120 minutes for protein measurements of GSH and TTase activity and for reverse transcription-polymerase chain reaction (RT-PCR) or Northern hybridization analysis to quantify TTase mRNA. The effect of GSH depletion on TTase mRNA expression was examined by treating the cells with buthionine S,R-sulfoximine (BSO); 1-chloro, 2,4-dinitrobenzene (CDNB); or 1,3-bis (2-chloroethyl)-1-nitrosourea (BCNU). Lens epithelial cells, depleted of cellular GSH by treatment with BCNU, were subjected to oxidative stress to examine the effect on TTase activity and mRNA level.

RESULTS

A transient increase was detected in TTase mRNA after 5 minutes of H2O2 treatment. The upregulation reached a maximum of 80% above the normal level by 10 minutes and gradually decreased as the oxidant was detoxified by the cells. Manipulation of cellular GSH level by treatment with BSO, CDNB, and BCNU resulted in a minimum change in TTase expression. It is noteworthy that when cells depleted of GSH were subjected to oxidative stress, TTase expression was also found to be strongly upregulated.

CONCLUSIONS

These observations suggest that the upregulation of TTase expression in the lens epithelial cells could be an adaptive response of the cells to combat oxidative stress to restore the vital functions of the lens proteins and enzymes. Such regulation is independent of cellular GSH concentration.

Authors+Show Affiliations

Department of Veterinary and Biomedical Sciences, University of Nebraska, Lincoln 68583-0905, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

11274078

Citation

Raghavachari, N, et al. "Regulation of Thioltransferase Expression in Human Lens Epithelial Cells." Investigative Ophthalmology & Visual Science, vol. 42, no. 5, 2001, pp. 1002-8.
Raghavachari N, Krysan K, Xing K, et al. Regulation of thioltransferase expression in human lens epithelial cells. Invest Ophthalmol Vis Sci. 2001;42(5):1002-8.
Raghavachari, N., Krysan, K., Xing, K., & Lou, M. F. (2001). Regulation of thioltransferase expression in human lens epithelial cells. Investigative Ophthalmology & Visual Science, 42(5), 1002-8.
Raghavachari N, et al. Regulation of Thioltransferase Expression in Human Lens Epithelial Cells. Invest Ophthalmol Vis Sci. 2001;42(5):1002-8. PubMed PMID: 11274078.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Regulation of thioltransferase expression in human lens epithelial cells. AU - Raghavachari,N, AU - Krysan,K, AU - Xing,K, AU - Lou,M F, PY - 2001/3/29/pubmed PY - 2001/5/18/medline PY - 2001/3/29/entrez SP - 1002 EP - 8 JF - Investigative ophthalmology & visual science JO - Invest Ophthalmol Vis Sci VL - 42 IS - 5 N2 - PURPOSE: To study how the expression of thioltransferase (TTase), a critical thiol repair and dethiolating enzyme, is regulated in human lens epithelial cells under oxidative stress. Also to examine whether depleting the primary cellular antioxidant glutathione (GSH) in these cells has any influence on TTase expression under the same conditions. METHODS: Human lens epithelial cells (B3) were grown to confluence (1.6 million) and gradually weaned from serum in the medium before exposing to 0.1 mM H2O2 for 2 hours. Cells were removed at the time intervals of 0, 5, 10, 15, 30, 60, and 120 minutes for protein measurements of GSH and TTase activity and for reverse transcription-polymerase chain reaction (RT-PCR) or Northern hybridization analysis to quantify TTase mRNA. The effect of GSH depletion on TTase mRNA expression was examined by treating the cells with buthionine S,R-sulfoximine (BSO); 1-chloro, 2,4-dinitrobenzene (CDNB); or 1,3-bis (2-chloroethyl)-1-nitrosourea (BCNU). Lens epithelial cells, depleted of cellular GSH by treatment with BCNU, were subjected to oxidative stress to examine the effect on TTase activity and mRNA level. RESULTS: A transient increase was detected in TTase mRNA after 5 minutes of H2O2 treatment. The upregulation reached a maximum of 80% above the normal level by 10 minutes and gradually decreased as the oxidant was detoxified by the cells. Manipulation of cellular GSH level by treatment with BSO, CDNB, and BCNU resulted in a minimum change in TTase expression. It is noteworthy that when cells depleted of GSH were subjected to oxidative stress, TTase expression was also found to be strongly upregulated. CONCLUSIONS: These observations suggest that the upregulation of TTase expression in the lens epithelial cells could be an adaptive response of the cells to combat oxidative stress to restore the vital functions of the lens proteins and enzymes. Such regulation is independent of cellular GSH concentration. SN - 0146-0404 UR - https://www.unboundmedicine.com/medline/citation/11274078/Regulation_of_thioltransferase_expression_in_human_lens_epithelial_cells_ L2 - https://iovs.arvojournals.org/article.aspx?volume=42&issue=5&page=1002 DB - PRIME DP - Unbound Medicine ER -