Fig. 4.

Overexpression of Gpc-1 and supplementation with arginine (Arg) increase the capacity to generate AM-containing heparan sulfate (HS-anMan) in SH-SY5Y cells. Representative immunofluorescence images of SH-SY5Y cells, which were treated with lipofectamine without vector (A, C, mock) or with a vector encoding green fluorescent protein (GFP)-tagged mouse Gpc-1 (B, D) in the absence (A, B) or presence of 1 mM Arg (C, D) or left untreated (E, UT) or exposed to 1 mM sodium nitroprusside (SNP) for 30 min (F). Staining was performed with mAb AM (for HS-anMan, A-D, red; Alexa Fluor 594-tagged secondary antibody), mAb S1 (for Gpc-1-SNO, E-F, green), anti-human Gpc-1 (E-F, red) and DAPI (for nuclei, blue), while GFP-Gpc-1 is visualized in the green channel of A-D. The intensities of HS-anMan and DAPI staining in A-D were measured and expressed as the ratio AM/DAPI. Exposure time was the same in all cases. Bar, 20 μm.