Minocycline blocks the activation of microglia; attenuates Pb-induced secrete of TNF-α, IL-1β and expression of iNOS; and protects co-cultured hippocampal neurons.

<p>Purified microglia were treated with vehicle (control) (A), Pb (50 µmol) (B), or Pb (50 µmol)+minocycline (45 µmol) (C) for 48 h, and were co-cultured with hippocampal neurons for another 48 h. (A, B, C) Microglia activation was detected by OX-42 antibody. (D) The results were quantified. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043924#s3" target="_blank">Results</a> (A, B and C) are expressed as the mean ± S.D. of average activated cell rate in random fields (n = 20). **<i>P</i><0.001 compared with control groups. *<i>P</i><0.01 compared with Pb-treated groups. iNOS was detected by anti-iNOS antibody (E, F, G. red fluorescence). Activation of iNOS was evaluated by measure the average fluorescence of iNOS-immunoreactive cells. (H) The results were quantified. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043924#s3" target="_blank">Results</a> (E, F and G) are expressed as the mean ± S.D. of average fluorescence of iNOS-immunoreactive cell number in random fields (n = 20). **<i>P</i><0.001 compared with control groups. *<i>P</i><0.01 compared with Pb-treated groups. TNF-α (I) and IL-1β (J) were determined by ELISA. Purified microglia were treated with vehicle (control) or Pb (50 µmol), or Pb (50 µmol)+minocycline (45 µmol) for 48 h for the TNF-α and IL-1β assay, respectively. **<i>P</i><0.001 compared with control groups. *<i>P</i><0.01 compared with Pb-treated groups. Apoptotic hippocampal neurons were detected and compared using TUNEL (green fluorescence) (L–T). Degeneration of hippocampal neurons was evaluated by counting the number of apoptotic neurons. (K) The results were quantified and are expressed as the mean ± S.D. of apoptotic neurons percentage in random fields (n = 20). **<i>P</i><0.001 compared with control groups. *<i>P</i><0.01 compared with Pb-treated groups. All scale bar indicates 50 µm.