Fig. 3. CaBP-9k knockout induces ER stress-induced apoptosis. (A) Western blotting with antibodies for ER stress-related proteins using brain lysates from old wild-type and CaBP-9k KO mice. (B) Quantification of A. n = 8 for mice for each group. The apoptosis markers were increased in old CaBP-9k KO mice. (C) Western blotting with antibodies for apoptosis-related proteins using brain lysates from old wild-type and CaBP-9k KO mice. (D) Quantification of C. n = 8 for mice for each group. The intensities of the protein bands were normalized to the GAPDH level. (E,G) Cell death was assessed in the hippocampi of old wild-type and CaBP-9k KO mice by immunostaining for cleaved caspase-3 or by TUNEL assays. Scale bar= 50 µm. (F,H) Quantification of E and G. n = 4 for mice for each group. (I) Immunostaining of hippocampal sections from old CaBP-9k KO mice for the proliferation marker Ki67. Scale bar= 100 µm. (J) Quantification of I. n = 4 for mice for each group. (K) Primary neuronal cells were cultured from embryonic day 13.5 wild-type and CaBP-9k KO mice for 2 days and then incubated with 100 µM BrdU for 24 h before immunostaining for Ki67 and BrdU. Scale bar= 40 µm. (L,M) Quantification of K. n = 5 cell culture replicates using 5 mice for each condition. (N) Primary neuronal cells cultured from embryonic day 12.5 wild-type and CaBP-9k KO mice for 3 days were transfected with pSuper_GFP/shCaBP-9k construct and then treated with BrdU for 24 h before immunostaining for GFP and BrdU. Scale bar= 40 µm. (O) Quantification of N. n = 7 cell culture replicates using 7 mice for each condition. Data shown are the means ± SEMs and were analysed by two-tailed unpaired Student's t-tests.