Objective: To investigate whether nuclear factor-kappaB (NF-kappaB) contributes to passively sensitized human airway smooth muscle cell (HASMCs) proliferation, and whether it is the downstream factor of activated protein kinase C (PKC).
Methods: HASMCs in culture were passively sensitized with 10% serum from asthmatic patients, with non-asthmatic human serum treated HASMCs as the control. NF-kappaB specific inhibitor pyrrolidine dithiocarbamate (PDTC) and PKC agonist phorbol 12-myristate 13-acetate (PMA) were used to intervene HASMCs exposed to asthmatic serum and non-asthmatic control serum. The proliferation of HASMCs was examined by cell cycle analysis, MTT colorimetric assay and proliferating cell nuclear antigen (PCNA) immunofluorescence staining respectively. NF-kappaB activity was detected by NF-kappaBp65 immunofluorescence staining and electrophoretic mobility shift assay (EMSA) respectively.
Results: (1) The percentage of S phase, A value, the positive expression rate of PCNA, the positive expression rate of NF-kappaBp65 and EMSA value in HASMCs passively sensitized with asthmatic serum were (21.78 +/- 2.79)%, 0.466 +/- 0.058, (67.5 +/- 8.5)%, (12.6 +/- 2.2)% and 32 781 +/- 9499 respectively. They were significantly increased compared with those of the control serum group (P < 0.05). After previously treated with PDTC, the above figures were decreased to (16.37 +/- 3.05)%, 0.389 +/- 0.035, (53.4 +/- 5.1)%, (4.9 +/- 1.3)% and 3934 +/- 937 respectively (P < 0.05). (2) After HASMCs were treated with both PMA and asthmatic serum, the percentage of S phase, A value, the positive expression rate of PCNA, the positive expression rate of NF-kappaBp65 and EMSA value were (25.52 +/- 3.38)%, 0.572 +/- 0.054, (81.2 +/- 10.2)%, (26.5 +/- 5.0)% and 71 654 +/- 12 293 respectively. After previously treated with PDTC, the above figures were (16.42 +/- 2.72)%, 0.386 +/- 0.031, (54.2 +/- 5.3)%, (5.9 +/- 1.4)% and 4808 +/- 1084 respectively. The difference was significant (P < 0.05).
Conclusions: NF-kappaB may contribute to the proliferation of HASMCs passively sensitized with human asthmatic serum, which involves the PKC/NF-kappaB signal pathway.