Figure 3. CARD6 knockout enhances apoptosis after SCI. (A) Representative images of NeuN/TUNEL double staining in dorsal horn of mice. The number of apoptotic cells (TUNEL positive) was quantified. Scale bar: 100 μm. (B, C) Western blot analysis of Bcl-2, Bax and cleaved Caspase-3 protein expression levels in the lumbar spinal cord segments. (D) Western blot analysis of mitochondrial and whole cell Cyto-c in the lumbar spinal cord segments. (E, F) BV2 cells with or without CARD6 knockdown were treated with LPS (100 ng/ml) for 24 h. (E) Then, IF staining was used to determine Iba1, Cyto-c and TUNEL levels. The quantification of Iba1, Cyto-c and TUNEL expression levels was exhibited. Scale bar: 50 μm. (F) Western blot analysis was used to calculate Cyto-c protein levels in mitochondria, or Cyto-c, Bcl-2, Bax and cleaved Caspase-3 expression in whole cell as indicated. Data represented means ± SEM (n=8 each group for in vivo studies; n=6 each group for in vitro studies). *p < 0.05 and **p < 0.01.