Figure 5

Inhibitors of HSPG maturation have no effect on adipocyte differentiation. 3T3-L1 pre-adipocytes were incubated with or without 3 mM pNP-Xyl or 4-MUmb concurrently with differentiation reagents for 4 d. Media was then changed to include just insulin with or without 3 mM pNP-Xyl or 4-MUmb for an additional 4–8 d. (A), cell lysates were immunoblotted with anti-VLDL-R mAb (1:50, culture supernatant), anti-LRP antisera (1:2000), or anti-CD36 IgG (5 μg/ml). For anti-leptin, proteins in culture supernatant were concentrated 10-fold by acetone precipitation prior to SDS-PAGE fractionation and immunoblotted with anti-leptin IgG (2 μg/ml). (B), following differentiation and xyloside treatment, cells were cultured for 16 h in serum-free, phenol red-free media. Culture supernatants were then assayed for LPL enzymatic activity as described in Methods. No significant difference was found in LPL levels between adipocytes treated with xylosides and untreated adipocytes (asterisk, p-value <0.01).