Figure 1.

T cell activation led to increased cell size and phospholipid levels as well as upregulated SELENOI. CD3⁺ T cells purified from mouse spleens were unstimulated or activated with anti-CD3/CD28 for 18 h. (A) Activated T cells showed increased sizes (forward scatter) using flow cytometry. Scale bar = 30 mm. (B) LCMS was used to compare levels of different lipid species extracted from T cells (displayed at pmol/10⁶ cells) and changes in PEs, plasmenyl PEs, and PCs are shown. (C) Real-time PCR was used to analyze mRNAs for selenoproteins and selenoprotein synthesis factors in unstimulated and activated T cells. Target mRNAs were normalized to housekeeping mRNA (ubc), β-actin was included as a comparison control, and il2ra (encodes for CD25) was included as a positive control. (D–E) Representative western blotting and densitometry analysis of SELENOI protein levels in 3 biological replicate experiments showed increases upon activation, and β-actin was used as a loading control. (F) Thin layer chromatography was used to analyze SELENOI activity (left) and the results of three independent experiments showed increases upon T cell activation (right). (G–I) Human T cells isolated from peripheral blood exhibited increases in SELENOI mRNA and protein after 18 h of anti-CD3/CD28 activation. Target mRNAs were normalized to housekeeping mRNA (ubc), β-actin was included as a comparison control, and CD25 was included as a positive control. Means of biological replicates (N = 3) were compared using Student's t test and expressed as mean ± SEM with ∗p < 0.05.