Skip to main content
. 2012 Mar 23;7(3):e32989. doi: 10.1371/journal.pone.0032989

Figure 5. Hypoxia-induced downregulation of PEDF in melanocytes and SBcl2 melanoma cells is not mediated by metalloproteinases or proteasomal degradation.

Figure 5

(A) Western blot analysis of extracellular PEDF (PEDFe) protein levels in 24 h conditioned medium (CM) from M330 primary melanocytes (upper blot) and SBcl2 melanoma cell line (lower blot). Cells were treated with 1 mM DMOG for 24 h and the CM were incubated with 100 ng human recombinant PEDF (rhuPEDF) and 20 mM EDTA at 37°C for 2 h. (B) Western blot analysis of PEDFe protein levels in CM from SBcl2 melanoma cell line treated with metalloproteinase inhibitor GM6001 (10 µM) and incubated under normoxia (21% O2) or hypoxia (1% O2) for 24 h. (C) Western blot analysis of PEDFe protein levels in 16 h CM and HIF1α protein levels in whole-cell extracts from SBcl2 and WM164 melanoma cell lines after treatment with the proteasome inhibitor MG132 (5 µM and 1 µM respectively) under normoxia or hypoxia. β-tubulin was used as loading control.