Fig. 5

In a human antibody-mediated CDC model, TNF-α promoted the cytotoxicity of PIECs and was dependent on Occludin. a-c PIECs were infected with control lentivirus or with lentivirus expressing Zo 1- (a), Claudin 2- (b), or Occludin- (c) specific siRNA. After 4 days, total RNA was collected and the mRNA level of Zo 1 (a), Claudin 2 (b), or Occludin (c) was analysed by RT-PCR (left side), and lysates were analyzed by western-blotting (right side) with antibodies against Zo 1 (a), Claudin 2 (b), Occludin (c), or actin. d-f The infected cells shown in (a-c), which corresponded to (d-f) respectively, were treated with or without rhTNF-α (20 ng/ml) for 48 h, and then exposed to human serum to induce antibody-mediated CDC. g PIECs infected with control virus (EV) or retrovirus encoding Occludin were left untreated or treated with rhTNF-α (20 ng/ml) for 48 h. Lysates were analyzed by immunoblotting with antibodies against Occludin and actin. h The infected cells shown in (g) were treated with or without rhTNF-α (20 ng/ml) for 48 h and then exposed to human serum to induce antibody-mediated CDC. Data are representative of at least three independent experiments (mean ± SEM). *p < 0.05, **p < 0.01, ***p < 0.001 by Student’s t test. N.S. = no significance, L.E. = long exposure, S.E. = short exposure