Fig. 2

Inhibition of proteasome suppresses microglial DICER downregulation induced by MPP⁺. a, b Quantitative PCR analysis of DICER mRNA levels in microglia (a, n = 4) and BV2 cells (b, n = 4) treated with MPP⁺ (100 μM) for 12 h. c Quantitative PCR examination of DICER mRNA levels in microglia sorted from VM of mice after saline or MPTP injection (n = 4). d Representative immunoblots of total lysates from microglia treated with MPP⁺ for the indicated times with the antibody against α-spectrin. e Representative immunoblots of total lysates from BV2 cells incubated with MPP⁺ together with the calpeptin (CAL, 10 μM) or MDL28170 (MDL, 60 μM) for 12 h and probed with the indicated antibodies. f Representative immunoblots of total lysates from microglia treated with MPP⁺ for the indicated times using the antibodies against caspase-3 or cleaved caspase-3. g Left—representative immunoblots of total lysates from BV2 cells incubated MPP⁺ with/without Ac-DEVD-CHO (DEVD, 10 μM) and probed with the indicated antibodies. Right—statistics (n = 4). h, i Upper—representative immunoblots of total lysates from microglia treated with MPP⁺ with/without lactacystin (LAC, 1 μM, h) or N-carbobenzoxyl-l-leucyl-l-leucyl-norvalinal (MG115, 1 μM, i) for 12 h and probed with the indicated antibodies. Lower—statistics (n = 3 for LAC and n = 4 for MG115). j Representative immunoblots of immunoprecipitates of total lysates from microglia treated with/without MPP⁺ in the presence of MG115 and then probed with anti-ubiquitin antibody. Unless stated, 100 μM MPP⁺ was used. The corresponding mRNA levels were normalized to actin levels. Data are mean + SEM. *p < 0.05, **p < 0.01, ***p < 0.001