Figure 5

A: Western blot detecting iNOS in PC12 cells simultaneously treated with 10 ng/ml NGF and 10 ng/ml TNFα for 24 hr. Thirty minutes before NGF/ TNFα treatment cells were pre-treated with 10 μM pyrrolidinedithyocarbamate (PDTC), 2 μM of a oligopeptide proteosome inhibitor (PSI) or 10 μM of a MAPK inhibitor (PD98059). Results shown are representative of 2 replicate experiments. B: SEAP release in the culture medium of PC12 cells transfected for 24 hr with an NF-κB-sensitive SEAP reporter gene construct and treated for 12 hr with vehicle (Control), 10 ng/ml NGF, 10 ng/ml TNFα or NGF plus TNFα in the presence of 10 μM PD98059, 10 μM PDTC or 2 μM PSI. Data are shown as mean ± S.E.M. from 3 independent replicate experiments. * and #: p < 0.05 vs. control or TNFα-alone cells, respectively (two-tailed unpaired Student's t-test). C: Representative photomicrographs of PC12 cells treated for 48 hr with 10 ng/ml NGF in the presence or absence of 10 μM PD98059 or 2 μM PDTC. D: NFκB transcriptional activity (as measured by a transiently transfected SEAP reporter vector) in PC12 cells treated for 24 hr with 10 ng/ml NGF, 10 ng/ml TNFα or NGF plus TNFα (Both) in the presence of 0.5 μM L-NAME. Data are shown as mean ± S.E.M. from 3 independent replicate experiments. * and #: p < 0.05 vs. control or TNFα-alone cells, respectively (two-tailed unpaired Student's t-test).