OBJECTIVES: The transcription factor NF-kB is involved in the expression of several genes linked to the immune response, including those of pro-inflammatory cytokines. We investigated cytokine production and NF-kB expression following CTLA4-Ig (abatacept) treatment of cultured human macrophages. METHODS: Human THP1 cells, differentiated in macrophages, were treated with CTLA4-Ig (100, 500 μg/ml; 3,12,24 hours). Quantitative RT-PCR analysis (qRT-PCR) of mRNA for NF-kB, IKBα and for IL-6, TNF-α, IL-1β, was performed after 3 and 12 hours from treatment. Western blot (WB) analysis for NF-kB and IKBα was performed after 24 hours from treatment. RESULTS: NF-kB gene expression was significantly downregulated (p<0.05), at 3 and 12 hours from CTLA4-Ig treatment, vs. untreated cells (cnt). IKBα resulted significantly increased vs. cnt (p<0.05), at 12 hours from CTLA4-Ig [500 μg/ml] treatment. After 3 hours, CTLA4-Ig [100 μg/ml] induced a significant decrease of TNF-α and IL-6 (p<0.05), vs. cnt and CTLA4-Ig [500 μg/ml] further reduced TNF-α (p<0.001), vs. cnt. After 12 hours from CTLA4-Ig treatment, a significant downregulation for IL-6 and IL1β expression (p<0.001), vs. cnt, was still evident. Results were confirmed by WB. CONCLUSIONS: NF-kB pathway seems to be implicated in the CTLA4-Ig modulation of macrophage cytokine expression. NF-kB expression resulted downregulated while its cytoplasmatic inhibitor IKBα was increased.
OBJECTIVES: The transcription factor NF-kB is involved in the expression of several genes linked to the immune response, including those of pro-inflammatory cytokines. We investigated cytokine production and NF-kB expression following CTLA4-Ig (abatacept) treatment of cultured human macrophages. METHODS:Human THP1 cells, differentiated in macrophages, were treated with CTLA4-Ig (100, 500 μg/ml; 3,12,24 hours). Quantitative RT-PCR analysis (qRT-PCR) of mRNA for NF-kB, IKBα and for IL-6, TNF-α, IL-1β, was performed after 3 and 12 hours from treatment. Western blot (WB) analysis for NF-kB and IKBα was performed after 24 hours from treatment. RESULTS: NF-kB gene expression was significantly downregulated (p<0.05), at 3 and 12 hours from CTLA4-Ig treatment, vs. untreated cells (cnt). IKBα resulted significantly increased vs. cnt (p<0.05), at 12 hours from CTLA4-Ig [500 μg/ml] treatment. After 3 hours, CTLA4-Ig [100 μg/ml] induced a significant decrease of TNF-α and IL-6 (p<0.05), vs. cnt and CTLA4-Ig [500 μg/ml] further reduced TNF-α (p<0.001), vs. cnt. After 12 hours from CTLA4-Ig treatment, a significant downregulation for IL-6 and IL1β expression (p<0.001), vs. cnt, was still evident. Results were confirmed by WB. CONCLUSIONS: NF-kB pathway seems to be implicated in the CTLA4-Ig modulation of macrophage cytokine expression. NF-kB expression resulted downregulated while its cytoplasmatic inhibitor IKBα was increased.
Authors: Diahann T S L Jansen; Hanane el Bannoudi; Ramon Arens; Kim L L Habets; Marjolijn Hameetman; Tom W J Huizinga; Jeroen N Stoop; René E M Toes Journal: Arthritis Res Ther Date: 2015-08-20 Impact factor: 5.156