Figure 2.

OPN and CLU are upregulated in denervated distal nerves cells. a, RT-PCR analysis shows that OPN and CLU mRNA levels are significantly upregulated in denervated sciatic nerve 7 d following transection (7 d Tx) and crush (7 d crush). mRNA levels for both factors return toward baseline levels 1 month following crush (1 m crush). b, Western blot analysis was performed on protein isolated from distal sciatic nerve 0, 1, 3, 7, and 14 d, and 1, 3, and 6 months following sciatic nerve transection. OPN protein expression begins to increase at 3 d, peaks between 14 d and 1 month, and decreases toward baseline levels by 3 and 6 months. The secreted form of CLU (80 kDa), not intracellular form (60 kDa), is the predominant isoform expressed in denervated sciatic nerve. CLU protein levels begin to increase at 3 d, with peak levels occurring 1 month after denervation, expression also remains high with extended denervation after 3 and 6 months. c, d, The cellular sources of OPN and CLU are denervated Schwann cells. c, Immunolabeling for both OPN (green) and S100 (red) shows that OPN is present in SCs (c) and in some axons (c, arrows) in intact nerve. Immunoreactivity increases in SCs that were denervated for 14 d. d, Immunoreactivity for CLU (green) was detected in subsets of SCs (red) in intact nerve (arrowheads) and increased immunoreactivity was detected in SCs that were denervated for 7 d. Scale bars, 100 μm.