PMID- 20038977
OWN - NLM
STAT- MEDLINE
DCOM- 20100308
LR  - 20211020
IS  - 1090-0535 (Electronic)
IS  - 1090-0535 (Linking)
VI  - 15
DP  - 2009 Dec 25
TI  - Specific NFkappaB subunit activation and kinetics of cytokine induction in 
      adenoviral keratitis.
PG  - 2879-89
AB  - PURPOSE: Corneal inflammation associated with ocular adenoviral infection is 
      caused by leukocytic infiltration of the subepithelial stroma in response to 
      expression of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) 
      by infected corneal cells. We have shown that these two chemokines are activated 
      by the mitogen-activated protein kinases (MAPKs) extracellular signal-regulated 
      kinase (ERK) and p38 for IL-8, and Jun-terminal kinase (JNK) for MCP-1. It is 
      also well established that transcription of each of these chemokines is tightly 
      controlled by the nuclear factor kappa B (NFkappaB) transcription factor family. 
      Therefore, we sought to better understand the differential regulation of 
      chemokine expression by NFkappaB in adenoviral infection of the cornea. METHODS: 
      Primary keratocytes derived from human donor corneas were treated with signaling 
      inhibitors and small interfering RNA specific to MAPKs, and infected with 
      adenovirus for different time periods before analysis. Activation of specific 
      NFkappaB subunits was analyzed by western blot, confocal microscopy, 
      electromobility shift assay, and chromatin immunoprecipitation, and chemokine 
      expression was quantified by enzyme-linked immunosorbent assay. RESULTS: Upon 
      adenoviral infection, NFkappaB p65, p50, and cREL subunits translocate to the 
      nucleus. This translocation is blocked by inhibitors of specific MAPK signaling 
      pathways. Confocal microscopy showed that inhibitors of the p38, JNK, and ERK 
      pathways differentially inhibited NFkappaB nuclear translocation, while PP2, an 
      inhibitor of Src family kinases, completely inhibited NFkappaB nuclear 
      translocation. Western blot analysis revealed that activation of specific 
      NFkappaB subunits was time dependent following infection. Chromatin 
      immunoprecipitation experiments indicated that binding of NFkappaB p65 and p50 
      subunits to the IL-8 promoter upon viral infection was differentially reduced by 
      chemical inhibitors of MAPKs. Electromobility shift assay and luciferase assay 
      analysis revealed that transactivation of IL-8 occurred with binding by the 
      NFkappaB p65 homodimer or NFkappaB p65/p50 heterodimer as early as 1 h post 
      infection, whereas MCP-1 expression was dependent upon the NFkappaB cREL but not 
      the p65 subunit, and occurred 4 h after IL-8 induction. Finally, knockdown of 
      NFkappaB p65 by short interfering RNA abrogated IL-8 but not MCP-1 expression 
      after adenoviral infection. CONCLUSION: The kinetics of NFkappaB subunit 
      activation are partly responsible for the observed pattern of acute inflammation 
      in the adenoviral-infected cornea. MAPKs differentially regulate chemokine 
      expression in adenoviral keratitis by differential and time-dependent activation 
      of specific NFkappaB subunits.
FAU - Rajaiya, Jaya
AU  - Rajaiya J
AD  - Department of Ophthalmology, Howe Laboratory, Massachusetts Eye and Ear 
      Infirmary, Harvard Medical School, Boston, MA 02114, USA.
FAU - Sadeghi, Neda
AU  - Sadeghi N
FAU - Chodosh, James
AU  - Chodosh J
LA  - eng
GR  - P30 EY014104/EY/NEI NIH HHS/United States
GR  - R01 EY013124/EY/NEI NIH HHS/United States
GR  - EY013124/EY/NEI NIH HHS/United States
GR  - P30EY014104/EY/NEI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PT  - Research Support, Non-U.S. Gov't
DEP - 20091225
PL  - United States
TA  - Mol Vis
JT  - Molecular vision
JID - 9605351
RN  - 0 (Chemokine CCL2)
RN  - 0 (Interleukin-8)
RN  - 0 (NF-kappa B)
RN  - 0 (Protein Subunits)
RN  - 0 (Transcription Factor RelA)
RN  - EC 2.7.11.10 (I-kappa B Kinase)
SB  - IM
MH  - Adenoviridae/*physiology
MH  - Adenoviridae Infections/complications/enzymology
MH  - Cell Nucleus/metabolism
MH  - Chemokine CCL2/genetics/*metabolism
MH  - Chromatin Immunoprecipitation
MH  - Enzyme Activation
MH  - Humans
MH  - I-kappa B Kinase/metabolism
MH  - Interleukin-8/genetics/*metabolism
MH  - Keratitis/complications/enzymology/*metabolism/*virology
MH  - Kinetics
MH  - NF-kappa B/antagonists & inhibitors/*metabolism
MH  - Promoter Regions, Genetic/genetics
MH  - Protein Binding
MH  - Protein Subunits/*metabolism
MH  - Protein Transport
MH  - Time Factors
MH  - Transcription Factor RelA/metabolism
PMC - PMC2797044
EDAT- 2009/12/30 06:00
MHDA- 2010/03/10 06:00
PMCR- 2009/01/01
CRDT- 2009/12/30 06:00
PHST- 2009/10/13 00:00 [received]
PHST- 2009/12/18 00:00 [accepted]
PHST- 2009/12/30 06:00 [entrez]
PHST- 2009/12/30 06:00 [pubmed]
PHST- 2010/03/10 06:00 [medline]
PHST- 2009/01/01 00:00 [pmc-release]
AID - 304 [pii]
AID - 2009MOLVIS0357 [pii]
PST - epublish
SO  - Mol Vis. 2009 Dec 25;15:2879-89.