A shed form of LDL receptor–related protein–1 regulates peripheral nerve injury and neuropathic pain in rodents
J. Clin. Invest. Alban Gaultier, et al. 118:161 doi:10.1172/JCI32371 [Go to this article.]

Figure 5
sLRP-α binds to Schwann cells in vitro and partially inhibits TNF-α binding. (A) Schwann cells were treated with FITC-labeled sLRP-α (green) or vehicle for 10 min at 37°C. The cells were washed, fixed, and visualized by fluorescence microscopy. Cell nuclei were stained by DAPI (blue). (B) Schwann cells were treated with 50 nM FITC-labeled sLRP-α (sLRP-α–FITC), 50 nM FITC-labeled IgG (IgG-FITC), or vehicle (unstained) for 1 h at 4°C. In some cases FITC-labeled sLRP-α was preincubated with 100 nM GST (sLRP-α–FITC+GST) or 100 nM GST-RAP (sLRP-α–FITC+GST-RAP) for 15 min at 22°C prior to incubation with cells. The cells were then washed and subjected to FACS. (C) Schwann cells were incubated with 50 nM sLRP-α for 15 min at 22°C. In some cases, sLRP-α was preincubated with 100 nM GST-RAP (sLRP-α+RAP) for 30 min at 37°C prior to incubation with the cells. Biotin-labeled TNF-α or vehicle (unstained) was then incubated with the cells for 1 h at 4°C. SBTI was incubated with separate cells as a negative control. FITC-labeled streptavidin was added for 30 min at 4°C. The cells were then washed and subjected to FACS. (D) Schwann cells were incubated with sLRP-α (+) or vehicle (–) for 15 min at 22°C. Biotin-labeled recombinant human TNF-α was added to some cells for 1 h at 4°C, followed by FITC-labeled streptavidin. Cells were washed and subjected to FACS. Data was analyzed using the FlowJo software, and the geometric mean of FITC fluorescence intensity is plotted. *P < 0.05 for reduction in TNF-α binding to Schwann cells compared with vehicle treatment (5–6 independent experiments).