ABSTRACT
Objectives To explore the effect of NF-?B signaling pathway on multicellular resistance(MCR) to 5-Fu in colon carcinoma cells.Methods The multicellular spheroids(MCS) of colon carcinoma HT-29 cell line were cultured by using liquid overlay technique,and the monolayer cells by using routine procedure.The morphology of MCS was observed with light microscope and scanning electronmicroscope(SEM).The expression of proliferating cell nuclear antigen(PCNA) was determined by immunohistochemistry.24 hours after adding 50?g/ml SN50 to spheroids cells,the 50% inhibiting concentration(IC50) for 5-Fu was determined by using radial outgrowth assay.The activity of NF-?B was measured by using electrophoretic mobility shift assay(EMSA) and the specific bands were observed.Apoptosis of spheroids cells,which were pretreated with 5?g/ml 5-Fu for 24 hours,was identified by TUNEL,the apoptosis related proteins caspase-3,Bcl-2 and Bax were detected by Western blotting and analyzed by semi-quantitative method.Results A necrotic core formed in the centre of MCS,and the peripheral cells were actively proliferating.Compared with the monolayer cultured cells,the activity of NF-?B was increased in MCS.When the activity of NF-?B in spheroid cells was inhibited by SN50,5-Fu readily induced cells to apoptosis(15.75%?1.02% vs 8.71%?0.73%,P