Figure 7
From: Sec-containing TrxR1 is essential for self-sufficiency of cells by control of glucose-derived H2O2
Low-glucose medium prevents cell death, Trx1 oxidation and JNK phosphorylation with Txnrd1−/− MEFs grown in sparse cell cultures. (a) Cells were seeded at a low density of 1000 cells/cm2 in either high-glucose (25 mM glucose, HG) or low-glucose (5.5 mM, LG) medium, whereupon they were cultured for either 24 or 96 h. Cell proliferation was then estimated by determination of total cellular nucleic acids (n=6–9, ±S.E.M.). Significant differences between the 96 h of Txnrd1−/− and the other 96 h samples are indicated (**P<0.01; ***P<0.001), and significant differences between the 24 and 96 h of each treatment are also indicated (#P<0.05; ##P<0.01; ###P<0.001; n.s., not significant, P>0.05). (b) Txnrd1fl/fl or Txnrd1−/− MEFs, with or without supplementation with catalase (100 units/ml), were cultured at 1000 cells/cm2 for 20 h in either HG (25 mM) or LG (5.5 mM) medium. The redox state of Trx1 was then analyzed by redox immunoblotting.30 The migration of Trx1 in a standard curve of MEF protein lysate treated so that Trx1 variants in all forms from 0 to 6 free Cys thiol groups being present30 is shown in the first lane. (c) The same set up as in (b) was used but phosphorylated JNK was determined by immunoblotting, using GAPDH as loading control
