Figure 4

Thesplenocyte apoptosis induced through E. adenophorum was mediated through the mitochondrial pathway.

(A) E. adenophorum treatment did not promote the cleavage of Bid but did increase the protein levels of Cytc and Bax and decreased the protein level of Bcl-2. The concentrations of Bax and Bcl-2 in splenocytesweremeasured through ELISA. The data are presented as the means ± SD of three independent experiments. *p < 0.05 and **p < 0.01 compared with the control group. (B) E. adenophorum treatment decreased the relative mRNA level of Bcl-2 but increased the level of Bax, resulting in an increased ratio of Bax/Bcl-2. (C) The protein levels of Fas or FasL were measured through western blotting and the results did not reveal any changes. (D)The splenocytes were subjected to western blot analysis to detect total and activated PARP. (E) E. adenophorum induced Bax translocation and Cytc release. The proteins in the cytosolic and mitochondrial fraction were collected and subsequently detected through western blotting. COX IV and β-actin were used as internal controls for the mitochondrial and cytosolic fractions, respectively. (F) E. adenophorum treatment induced collapse of ΔΨm. The cell suspension was filtered through 300-mesh nylon and then stained with JC-1, followed by FCM analysis.