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. 2008 Nov 5;83(2):598–611. doi: 10.1128/JVI.00629-08

FIG. 2.

FIG. 2.

MnSOD is induced in vFLIP/K13-expressing HUVEC and in HHV-8-infected HUVEC. (A) Protein lysates from Ctrl-EC or K13-EC were labeled with either Cy5 (Ctrl-EC) or Cy3 (K13-EC). The lysates were subjected to 2D-DIGE. Signals were detected by laser scanning and quantified with analysis software. The spot with the highest surplus in K13-EC-derived extracts (arrow) was isolated after Coomassie staining and subjected to mass spectroscopic analysis (Toplab GmbH, Marinsried, Germany). This identified MnSOD. (B) Western blot analysis confirmed increased MnSOD expression in K13-EC. vFLIP/K13 expression was detected with an anti-Myc antibody; MnSOD and GAPDH were detected with specific antibodies directed against the proteins. (C) Immunocytochemical staining of MnSOD in Ctrl-EC and K13-EC showed that MnSOD was selectively detectable in K13-transduced cells (red; arrowheads). The cells were counterstained by hematoxylin (light blue). (D) HUVEC were infected with a recombinant HHV-8 encoding GFP (HHV-8-EC). Phase-contrast (PhaCo) analysis of cell density and fluorescence microscopy (FM) for detection of HHV-8-infected cells confirmed efficient infection of HUVEC 72 h postinfection. After puromycin selection, the infected cells (HHV-8-EC) were subjected to Western blot analysis. Comparison of uninfected cells (HUVEC), Ctrl-EC, and K13-EC revealed upregulation of MnSOD in HHV-8-infected cells. GAPDH detection was used as a loading control.