Effects of Tanshinone IIA in Adult Rats after SCI Figure 8. Effects of TIIA on apoptosis in spinal cord after SCI. Few apoptotic cells in the tissues from the sham and sham/TIIA groups were observed (A–B). SCI induced a marked increase of apoptotic cells at 3 days after trauma (C). In contrast, tissues obtained from TIIA treatement group showed significantly less apoptotic cells (D). The number of TUNEL positive cells was counted in 5 to 10 fields for each slide and the analyzed data are shown (E). Bars represent means 6 SEM. **p,0.001 vs. sham group, #p,0.05 vs. SCI group. doi:10.1371/journal.pone.0038381.g008 Effects of TIIA on the expression of iNOS after SCI iNOS is another inflammation-related mediator. We evaluated the expression of iNOS at 24 h after SCI. The results of the real- time PCR analysis showed that SCI caused a significant increase in iNOS mRNA expression in the SCI group (Fig. 7A). However, this increase was markedly downregulated upon the intraperito- neal injection of TIIA in the SCI/TIIA group (Fig. 7A). In addition, the results of the western blot analysis indicated that the SCI-induced iNOS protein expression was also decreased in the SCI/TIIA group (Fig. 7B and C). Effects of TIIA on apoptosis in spinal cord after SCI To determine whether TIIA could affect SCI-induced apopto- sis, we performed TUNEL staining on spinal cord sections. We found that the highest amount of apoptotic cells occurred at Day 3 after SCI in our preliminary experiment, thus we chose to analyze Figure 9. Effects of TIIA on apoptosis-related factors after SCI. The protein expression of Bcl-2 was reduced in the SCI group at 1 day and 3 days compared to the sham and sham/TIIA groups, whereas Bcl-2 was up-regulated in the SCI/TIIA group (A). The analyzed data are shown. (B) The cleaved caspase-3 expression began to increase at 1 day, peaked at 3 days and quickly reduced to a lower level at 7 days after SCI. However, TIIA treatment decreased the cleaved caspase-3 (C). The analyzed data are shown (D). Bars represent means 6 SEM. *p,0.05 vs. sham group, #p,0.05 vs. SCI group. doi:10.1371/journal.pone.0038381.g009 PLoS ONE | www.plosone.org 7 June 2012 | Volume 7 | Issue 6 | e38381