Biochemistry (Moscow)

, Volume 75, Issue 2, pp 201–207

Selenoprotein W depletion in vitro might indicate that its main function is not as an antioxidative enzyme

  • Xiao-Long Wang
  • Chuan-Ping Yang
  • Kai Xu
  • Ou-Jv Qin
Article

DOI: 10.1134/S0006297910020100

Cite this article as:
Wang, XL., Yang, CP., Xu, K. et al. Biochemistry Moscow (2010) 75: 201. doi:10.1134/S0006297910020100

Abstract

Examination of the antioxidative homeostasis in skeletal muscle cells in the presence or absence of selenoprotein W (SelW) is necessary to understand the importance of SelW in the antioxidative system. Depletion of SelW by RNA interference was achieved by introducing a synthetic small interfering RNA into the mouse skeletal muscle cell line C2C12 (C3H). Transfectant screening was performed by real-time reverse transcription-PCR, Western blotting, flow cytometry, fluorescence staining, cell viability, and glutathione assays. SelW expression and mRNA levels were downregulated by 62.1 and 72.4%, respectively. In addition, acute cytotoxicity and an apoptosis rate of ∼36% in SelW-depleted cells demonstrated that RNA interference was successful. As compared with non-SelW-depleted cells, the enzyme activities of glutathione peroxidase, superoxide dismutase, and catalase and total antioxidative capability and glutathione level increased by 47.6, 103.0, 31.0, 205.6, and 30.0%, respectively (P < 0.05). Thus, SelW is important for the antioxidative system of muscle cells. Depletion of SelW, however, could be compensated by other intracellular antioxidative enzymes because oxidative stress was not the causative factor for apoptosis in SelW-depleted cells. Thus, the main function of SelW in muscle cells is not in the antioxidative system.

Key words

selenoprotein WfunctionRNA interference antioxidationmuscle cellin vitro

Abbreviations

CAT

catalase

FITC

fluorescein isothiocyanate

GPx

glutathione peroxidase

GSH

glutathione

MTT

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

PI

propidium iodide

SelW

selenoprotein W

siRNA

small interfering RNA

SOD

superoxide dismutases

T-AOC

total antioxidative capability

WST-1

2-(4-iodophenyl)-3-(4-nitro-phenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium, monosodium salt

Supplementary material

10541_2010_9172_MOESM1_ESM.pdf (254 kb)
Supplementary material, approximately 254 KB.

Copyright information

© Pleiades Publishing, Ltd. 2010

Authors and Affiliations

  • Xiao-Long Wang
    • 1
    • 2
  • Chuan-Ping Yang
    • 2
  • Kai Xu
    • 1
  • Ou-Jv Qin
    • 1
  1. 1.Biomedicine Division, Wildlife Resource CollegeNortheast Forestry University, Ministry of EducationHarbinP. R. China
  2. 2.Key Laboratory of Forest Tree Genetics Improvement and BiotechnologyMinistry of EducationHarbinP. R. China