Attenuation of NF-κB Signaling Response to UVB Light during Cellular Senescence

Abstract The ability of cells to adapt to environmental stresses undergoes a progressive reduction during aging. NF-κB-mediated signaling is a major defensive system against various environmental challenges. The aim of this study was to find out whether replicative senescence affects the response of the NF-κB signaling pathway to UVB light in human WI-38 and IMR-90 fibroblasts. The exposure of early passage fibroblasts to UVB light inhibited the proliferation and induced a flat phenotype similar to that observed in replicatively senescent fibroblasts not exposed to UVB light. The UVB radiation dose used (153 mJ/cm 2 ) did not induce apoptosis in either early or late passage WI-38 fibroblasts. UVB exposure induced a prominent activation of the NF-κB signaling pathway both in early and in late passage WI-38 and IMR-90 fibroblasts. Interestingly, the response to UVB light was significantly attenuated in late passage fibroblasts. This attenuation was most prominent in DNA binding activities of nuclear NF-κB complexes. Similar senescence-related attenuation was also observed in the DNA binding activities of nuclear AP-1 and Sp-1 factors after UVB treatment. Immunoblotting and -cytochemistry showed an increase in nuclear localization of p50 and p65 components of NF-κB complexes. Supershift experiments showed that the specific NF-κB complexes contain p50 and p65 protein components but not p52 and c-Rel proteins. Cytoplasmic IκBα showed a marked decrease at protein level but an increase in phosphorylation after UVB treatment. Transient transfection assays with TK5-CAT and TK10-CAT plasmids carrying NF-κB-responsive sites of the TNFα promoter were used to analyze the functional activity of the NF-κB complexes. Results showed that UVB exposure induced an increase in NF-κB-driven CAT expression both in early and in late passage fibroblasts though the response was significantly stronger in early passage fibroblasts. Our results show that the induction of NF-κB-mediated signaling by UVB light is highly attenuated in senescent fibroblasts. This attenuation may reduce the stress resistance during cellular senescence.