Fig. 3
Neutrophil elastase (NE) inhibition via sivelestat prevented the spinal cord injury (SCI)-induced modulation of angiopoietins (ANGPTs) and inhibited the expression of NE. a Molecular structure of sivelestat (i) and concentrations determined in plasma (ii), brain (iii), and spinal cord (iv) at several time points as described in the Methods section (n = 2/timepoint). Samples from sham, injured untreated, and injured sivelestat-treated animals were prepared DPI-1 as described in the Methods section. Representative images of immunohistochemistry for ANGPT-1 and rat endothelial cell antigen (RECA-1) (b) and ANGPT-2 and NE (e) at 1 day after SCI [3 fields/slide, n = 2–3/group (sham = 2, injury = 3 and sivelestat = 3)]. c Western blots of ANGPT-1, p-AKT, and AKT expression at 1 day after injury. Actin was used as internal controls for western blot [n = 2–3/group (sham = 2, injury = 3 and sivelestat = 3)]. Total RNA and spinal extracts from sham or injured untreated (Injury) or after sivelestat treatment were prepared 1 day after the injury. RT-PCR results of Ang-1 (d), Ang-2 (f) and N.E (g) expression 1 day after injury [n = 2–3/group (sham = 2, injury = 3 and sivelestat = 3) performed in triplicates]. GAPDH was used as internal controls for real-time quantitative reverse transcription–polymerase chain reaction. Data represent means ± S.E.M. ###p < 0.001 vs. sham group. *p < 0.05, **p < 0.01, ***p < 0.001 vs. Injury group