American Society of Hematology

Figure 6.

$Inhibitors of hGIIA that block the binding of hGIIA to vimentin and HSPGs. (A) hGIIA preincubated or not with DTT (100 mM) or heparin (500 μg/mL) for 30 minutes at RT was injected into 1 mL columns containing heparan sulfates as described in “Materials and methods.” The columns were then washed, and the eluate was analyzed for hGIIA by immunoblotting as described in the “Materials and methods.” (B) Indicated amounts of vimentin were spotted on PVDF membranes. hGIIA (0.1 μg/mL) was preincubated or not with 10 mM DTT or 10 μM LY311727 at RT for 30 minutes and then incubated with immobilized vimentin on PVDF membranes in the presence or the absence of DTT or LY311727. The membrane was then probed with the hGIIA antibody as described in “Materials and methods.” (C) hGIIA was preincubated or not with LY311727 (10 μM) for 30 minutes at RT and then incubated with control or Fas-treated cells in the presence or the absence of heparin (50 μg/mL). Cell suspensions were sequentially lysed, and proteins in the Hypo LB–soluble and Hypo LB–insoluble/Hyper LB–soluble fractions were separated by SDS-PAGE. The samples were transferred to a PVDF membrane and probed with the hGIIA antibody as described in “Materials and methods.”$

Inhibitors of hGIIA that block the binding of hGIIA to vimentin and HSPGs. (A) hGIIA preincubated or not with DTT (100 mM) or heparin (500 μg/mL) for 30 minutes at RT was injected into 1 mL columns containing heparan sulfates as described in “Materials and methods.” The columns were then washed, and the eluate was analyzed for hGIIA by immunoblotting as described in the “Materials and methods.” (B) Indicated amounts of vimentin were spotted on PVDF membranes. hGIIA (0.1 μg/mL) was preincubated or not with 10 mM DTT or 10 μM LY311727 at RT for 30 minutes and then incubated with immobilized vimentin on PVDF membranes in the presence or the absence of DTT or LY311727. The membrane was then probed with the hGIIA antibody as described in “Materials and methods.” (C) hGIIA was preincubated or not with LY311727 (10 μM) for 30 minutes at RT and then incubated with control or Fas-treated cells in the presence or the absence of heparin (50 μg/mL). Cell suspensions were sequentially lysed, and proteins in the Hypo LB–soluble and Hypo LB–insoluble/Hyper LB–soluble fractions were separated by SDS-PAGE. The samples were transferred to a PVDF membrane and probed with the hGIIA antibody as described in “Materials and methods.”

This Feature Is Available To Subscribers Only