title: Tumor necrosis factor-{alpha} suppresses angiotensinogen expression through formation of a p50/p50 homodimer in human renal proximal tubular cells.
abstract: Angiotensinogen (AGT) expression in renal proximal tubular cells (RPTCs) and intrarenal tumor necrosis factor-α (TNF-α) levels are increased in hypertension and renal diseases However, the contribution of TNF-α to AGT expression in RPTCs has not been established. Therefore, the objective of the present study was to determine influence of TNF-α on AGT expression in RPTCs. Human kidney-2 (HK-2) cells, immortalized human RPTCs, were treated with several concentrations of TNF-α up to 24 h. AGT mRNA and protein expression were evaluated by RT-PCR and ELISA, respectively. Activation of nuclear factor-κB (NF-κB) by TNF-α was evaluated by Western blot analysis, immunocytochemistry, and electrophoretic mobility shift assay (EMSA). TNF-α suppressed AGT mRNA expression in a dose- and time-dependent manner. Maximum AGT mRNA reduction was caused by 40 ng/ml of TNF-α (0.52 ± 0.09, ratio to control, at 24 h) and at 24 h (0.66 ± 0.05, ratio to control, by 10 ng/ml TNF-α). TNF-α reduced AGT protein accumulation in the medium between 8 and 24 h (0.62 ± 0.13 by 40 ng/ml TNF-α, ratio to control). TNF-α activated and induced translocalization of p50 and p65, which are NF-κB subunits. Elevated formation of p50/p65 and p50/p50 dimers by TNF-α were observed by EMSA and supershift assay. Gene silencing of p50, but not p65, attenuated the effect of TNF-α on reduction of AGT expression in RPTCs. These results indicate that TNF-α suppresses AGT expression through p50/p50 homodimer formation in human RPTCs, suggesting a possible counteracting mechanism that limits excessive intrarenal AGT production.
authors.affiliation: Department of Medicine and Physiology, Molecular Core in Hypertension and Renal Center of Excellence, Tulane Univ. Health Sciences Center, 1430 Tulane Ave., No. SL39/M720, New Orleans, LA 70112, USA.
authors.forename: Ryousuke
authors.initials: R
authors.lastname: Satou
authors.forename: Kayoko
authors.initials: K
authors.lastname: Miyata
authors.forename: Akemi
authors.initials: A
authors.lastname: Katsurada
authors.forename: L Gabriel
authors.initials: LG
authors.lastname: Navar
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kobori
mesh_terms: D000808:Angiotensinogen / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D052002:NF-kappa B p50 Subunit / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000235:genetics / Q000378:metabolism; D047889:Receptors, Tumor Necrosis Factor, Type II / Q000235:genetics / Q000378:metabolism; D051996:Transcription Factor RelA / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D052002:NF-kappa B p50 Subunit; D034741:RNA, Small Interfering; D047888:Receptors, Tumor Necrosis Factor, Type I; D047889:Receptors, Tumor Necrosis Factor, Type II; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000808:Angiotensinogen