PMID- 9950608
OWN - NLM
STAT- MEDLINE
DCOM- 19990210
LR  - 20161124
IS  - 0146-0404 (Print)
IS  - 0146-0404 (Linking)
VI  - 40
IP  - 2
DP  - 1999 Feb
TI  - Suppression of NF-kappaB-dependent proinflammatory gene expression in human RPE 
      cells by a proteasome inhibitor.
PG  - 477-86
AB  - PURPOSE: To determine whether nuclear transcription factor-kappaB (NF-kappaB) is 
      activated in human retinal pigment epithelial (hRPE) cells in response to 
      interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), or 
      interferon-gamma (IFN-gamma) alone or in combination and if so, whether 
      expression of proinflammatory genes induced by these agents can be blocked by a 
      proteasome inhibitor, MG-132, which inhibits the degradation of I kappaB, an 
      NF-kappaB inhibitor, thereby preventing nuclear translocation of NF-kappaB. 
      METHODS: Cultured hRPE were pretreated for 60 minutes with medium alone or medium 
      containing the proteasome inhibitor MG-132 (20 microM) and then exposed to 
      TNF-alpha (1.1 x 10(3) U/ml), IL-1beta (5 U/ml), or IFN-gamma (7.5 x 10(3) U/ml) 
      alone or in combination (TII). Nuclear translocation of NF-kappaB was determined 
      by fluorescence staining of the NF-kappaB Rel A (p65) subunit. Cytoplasmic I 
      kappaB protein was measured by western blot analysis. Nuclear extract binding to 
      kappaB DNA motifs was measured by electrophoretic mobility shift assay and 
      antibody supershift assay. Steady state mRNA expression of the chemokines 
      melanoma growth stimulating activity (MGSA)/gro-alpha, regulated on activation 
      normal T-cell expression and secreted (RANTES), and monocyte chemoattractant 
      protein (MCP-1), the cytokines IL-1beta and macrophage colony stimulating factor 
      (M-CSF) and intercellular adhesion molecule-1 (ICAM-1) was evaluated by 
      semiquantitative reverse transcription-polymerase chain reaction. Chemokine and 
      cytokine protein secretion was measured by enzyme-linked immunosorbent assay. 
      Cell-surface ICAM-1 expression was determined by flow cytometry. RESULTS: 
      TNF-alpha, IL-1beta, and TII but not IFN-gamma alone caused degradation of I 
      kappaB, Rel A nuclear translocation, and increased NF-kappaB DNA binding 
      activity, effects that were blocked by pretreatment with MG-132. MG-132 
      suppressed MGSA/gro-alpha, RANTES, MCP-1, IL-1beta, M-CSF, and ICAM-1 mRNA 
      expression and secreted RANTES, MCP-1, and M-CSF protein, and cell-surface ICAM-1 
      that were induced by IL-1beta, TNF-alpha, and TII. CONCLUSIONS: TNF-alpha, 
      IL-1beta, and TII induce expression of proinflammatory cytokines and ICAM-1 in 
      hRPE cells through an NF-kappaB-dependent signal transduction pathway. This 
      effect is blocked by MG-132, a proteasome inhibitor that prevents I kappaB 
      degradation. Inhibition of NF-kappaB may be a useful strategy to treat 
      proliferative vitreoretinopathy and uveitis, ocular diseases initiated and 
      perpetuated by cytokine activation.
FAU - Wang, X C
AU  - Wang XC
AD  - Department of Ophthalmology, Duke University Medical Center, Durham, North 
      Carolina, USA.
FAU - Jobin, C
AU  - Jobin C
FAU - Allen, J B
AU  - Allen JB
FAU - Roberts, W L
AU  - Roberts WL
FAU - Jaffe, G J
AU  - Jaffe GJ
LA  - eng
GR  - 5P30EYO5722-12/EY/NEI NIH HHS/United States
GR  - EY11364-01/EY/NEI NIH HHS/United States
GR  - EYO9106/EY/NEI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Invest Ophthalmol Vis Sci
JT  - Investigative ophthalmology & visual science
JID - 7703701
RN  - 0 (CXCL1 protein, human)
RN  - 0 (Chemokine CCL2)
RN  - 0 (Chemokine CCL5)
RN  - 0 (Chemokine CXCL1)
RN  - 0 (Chemokines, CXC)
RN  - 0 (Chemotactic Factors)
RN  - 0 (Cysteine Proteinase Inhibitors)
RN  - 0 (Cytokines)
RN  - 0 (DNA Primers)
RN  - 0 (Drug Combinations)
RN  - 0 (Growth Substances)
RN  - 0 (Intercellular Signaling Peptides and Proteins)
RN  - 0 (Leupeptins)
RN  - 0 (NF-kappa B)
RN  - 0 (RNA, Messenger)
RN  - 0 (Transcription Factor RelA)
RN  - 126547-89-5 (Intercellular Adhesion Molecule-1)
RN  - 81627-83-0 (Macrophage Colony-Stimulating Factor)
RN  - RF1P63GW3K (benzyloxycarbonylleucyl-leucyl-leucine aldehyde)
SB  - IM
MH  - Blotting, Western
MH  - Cells, Cultured
MH  - Chemokine CCL2/genetics/metabolism
MH  - Chemokine CCL5/genetics/metabolism
MH  - Chemokine CXCL1
MH  - *Chemokines, CXC
MH  - Chemotactic Factors/genetics/metabolism
MH  - Cysteine Proteinase Inhibitors/*pharmacology
MH  - Cytokines/pharmacology
MH  - DNA Primers/chemistry
MH  - Drug Combinations
MH  - Enzyme-Linked Immunosorbent Assay
MH  - Gene Expression/*drug effects
MH  - Growth Substances/genetics/metabolism
MH  - Humans
MH  - Intercellular Adhesion Molecule-1/genetics/metabolism
MH  - *Intercellular Signaling Peptides and Proteins
MH  - Leupeptins/*pharmacology
MH  - Macrophage Colony-Stimulating Factor/genetics/metabolism
MH  - NF-kappa B/*genetics/*metabolism
MH  - Pigment Epithelium of Eye/*drug effects/metabolism
MH  - RNA, Messenger/metabolism
MH  - Transcription Factor RelA
EDAT- 1999/02/09 00:00
MHDA- 1999/02/09 00:01
CRDT- 1999/02/09 00:00
PHST- 1999/02/09 00:00 [pubmed]
PHST- 1999/02/09 00:01 [medline]
PHST- 1999/02/09 00:00 [entrez]
PST - ppublish
SO  - Invest Ophthalmol Vis Sci. 1999 Feb;40(2):477-86.