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Expression of superoxide dismutase in whole lens prevents cataract formation.
Mol Vis. 2005 Oct 11; 11:853-8.MV

Abstract

PURPOSE

Oxidative damage is a major factor causing cataracts, which account for almost half of human blindness cases worldwide. In this study, we wished to determine if overexpression of superoxide dismutase (SOD) in intact lenses could prevent cataract formation induced by oxidative stress.

METHODS

Fresh, intact lenses from 6-week-old male/female Sprague Dawley rats were incubated with plasmid DNA encoding the human SOD1 (Cu/Zn-SOD) gene at 37 degrees C in a CO2 cell culture chamber with 95% air and 5% CO2. SOD1 expression was determined by western blotting and SOD enzyme activity. Lenses with or without overexpression of SOD1 were treated with H2O2 and cataract formation was examined. SOD1 regulation of protein kinase Cgamma (PKCgamma) was determined by PKCgamma enzyme activity assay. Intact lens gap junctions were determined by dye transfer assay.

RESULTS

In the lens overexpression system, SOD1 cDNA was fused to EYFP to generate EYFP:SOD1 fusion proteins which allow detection from endogenous SOD1. Incubation of intact lenses with plasmid DNA produced EYFP:SOD1 fusion proteins as determined by western blot using anti-GFP or anti-SOD1 antibodies. This caused significant increases in SOD enzyme activity. Data indicated that SOD1 plasmid DNA can be expressed as a functional enzyme in intact lenses in culture. Lenses overexpressing SOD1 remained clear after H2O2 treatment at 100 muM for 24 h, similar to control. Overexpression of SOD1 diminished the effect of H2O2 on PKCgamma activation and subsequent inhibition of gap junctions, indicating that overexpression of SOD1 may reduce reactive oxygen species (ROS) production, and this would prevent the normal H2O2 effect on cataract formation.

CONCLUSIONS

Overexpression of SOD1 in whole lens prevents H2O2-induced oxidative damage (cataract formation) to the lens and subsequent control of gap junctions by protein kinase Cgamma.

Authors+Show Affiliations

Department of Biochemistry, Kansas State University, Manhattan, KS 66506, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

16254550

Citation

Lin, Dingbo, et al. "Expression of Superoxide Dismutase in Whole Lens Prevents Cataract Formation." Molecular Vision, vol. 11, 2005, pp. 853-8.
Lin D, Barnett M, Grauer L, et al. Expression of superoxide dismutase in whole lens prevents cataract formation. Mol Vis. 2005;11:853-8.
Lin, D., Barnett, M., Grauer, L., Robben, J., Jewell, A., Takemoto, L., & Takemoto, D. J. (2005). Expression of superoxide dismutase in whole lens prevents cataract formation. Molecular Vision, 11, 853-8.
Lin D, et al. Expression of Superoxide Dismutase in Whole Lens Prevents Cataract Formation. Mol Vis. 2005 Oct 11;11:853-8. PubMed PMID: 16254550.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Expression of superoxide dismutase in whole lens prevents cataract formation. AU - Lin,Dingbo, AU - Barnett,Micheal, AU - Grauer,Laura, AU - Robben,Jerry, AU - Jewell,Annie, AU - Takemoto,Larry, AU - Takemoto,Dolores J, Y1 - 2005/10/11/ PY - 2005/10/29/pubmed PY - 2006/4/14/medline PY - 2005/10/29/entrez SP - 853 EP - 8 JF - Molecular vision JO - Mol Vis VL - 11 N2 - PURPOSE: Oxidative damage is a major factor causing cataracts, which account for almost half of human blindness cases worldwide. In this study, we wished to determine if overexpression of superoxide dismutase (SOD) in intact lenses could prevent cataract formation induced by oxidative stress. METHODS: Fresh, intact lenses from 6-week-old male/female Sprague Dawley rats were incubated with plasmid DNA encoding the human SOD1 (Cu/Zn-SOD) gene at 37 degrees C in a CO2 cell culture chamber with 95% air and 5% CO2. SOD1 expression was determined by western blotting and SOD enzyme activity. Lenses with or without overexpression of SOD1 were treated with H2O2 and cataract formation was examined. SOD1 regulation of protein kinase Cgamma (PKCgamma) was determined by PKCgamma enzyme activity assay. Intact lens gap junctions were determined by dye transfer assay. RESULTS: In the lens overexpression system, SOD1 cDNA was fused to EYFP to generate EYFP:SOD1 fusion proteins which allow detection from endogenous SOD1. Incubation of intact lenses with plasmid DNA produced EYFP:SOD1 fusion proteins as determined by western blot using anti-GFP or anti-SOD1 antibodies. This caused significant increases in SOD enzyme activity. Data indicated that SOD1 plasmid DNA can be expressed as a functional enzyme in intact lenses in culture. Lenses overexpressing SOD1 remained clear after H2O2 treatment at 100 muM for 24 h, similar to control. Overexpression of SOD1 diminished the effect of H2O2 on PKCgamma activation and subsequent inhibition of gap junctions, indicating that overexpression of SOD1 may reduce reactive oxygen species (ROS) production, and this would prevent the normal H2O2 effect on cataract formation. CONCLUSIONS: Overexpression of SOD1 in whole lens prevents H2O2-induced oxidative damage (cataract formation) to the lens and subsequent control of gap junctions by protein kinase Cgamma. SN - 1090-0535 UR - https://www.unboundmedicine.com/medline/citation/16254550/Expression_of_superoxide_dismutase_in_whole_lens_prevents_cataract_formation_ L2 - http://www.molvis.org/molvis/v11/a101/ DB - PRIME DP - Unbound Medicine ER -