Figure 6

Crosslinking of tyrHDL to ABCA1. Cholesterol-loaded fibroblasts were equilibrated in the presence of 5 μM T0901317 for 24 h to upregulate ABCA1 expression. The cells were then treated with BSA alone or plus 10 μg/ml apoA-I, HDL and tyrHDL for 2 h, washed, and then incubated with 250 mM of DSP for 1 h. Whole cell lysates were isolated, run on a 5-15% SDS-PAGE gradient gel and probed for the presence of ABCA1 and apoA-I by immunoblot. 5% beta mercaptoethanol (β-ME) was used as a reducing agent to cleave crosslinks formed by DSP. Results are representative of three separate experiments with similar results.