[Effect of mm-LDL on NF-kB activation in endothelial cell].
OBJECTIVE
To investigate the signal transduction pathway of NF-kB activated by minimally modified low density lipoprotein (mm-LDL) in endothelial cells and the effect of NF-kB on platelet derived growth factor b (PDGFb) mRNA expression.


METHODS
mm-LDL was prepared through iron oxidation by dialyzing the native LDL against FeSO4 in PBS. Endothelial cells were incubated in a medium containing mm-LDL, TNF, and IL-1 respectively and electrophoretic mobility shift assay (EMSA) was displayed to check on the activation of NF-kB. Luciferase reporter gene was analysed to investigate the effect of nuclear factor inducing kinase (NIK), inhibitor of NF-kB kinase alpha (IKK alpha) and inhibitor of NF-kB kinase beta (IKK beta) on NF-kB activation. In addition, endothelial cells were transfected using PDGFb promoter-luciferase for reporter gene analysis or transfected with mut-NIK for slot blot analysis to study the effect of NF-kB on PDGFb mRNA expression.


RESULTS
mm-LDL was able to activate NF-kB in endothelial cells. mut-NIK and mut-IKK beta inhibited luciferase activity induced by mm-LDL. mm-LDL could also enhance luciferase activity controlled by upstream sequence of PDGFb promoter which contains element interacting with NF-kB. Result of slot blot showed inhibition of PDGFb mRNA expression by mut-NIK in the endothelial cells stimulated by mm-LDL.


CONCLUSIONS
mm-LDL may activate NF-kB through NIK-IKK beta pathway and promote PDGFb mRNA expression in endothelial cells.
Yin
H C
HC
Department of Pathology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
Liu
X Y
XY
Liu
P M
PM
Zhang
H
H
Liang
P
P
Wang
Z L
ZL
She
M P
MP
D002478:Cells, Cultured; D024202:Electrophoretic Mobility Shift Assay; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D051550:I-kappa B Kinase; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D016328:NF-kappa B / Q000187:drug effects* / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D020574:Proto-Oncogene Proteins c-sis / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D015398:Signal Transduction
D008077:Lipoproteins, LDL; D016328:NF-kappa B; D020574:Proto-Oncogene Proteins c-sis; D012333:RNA, Messenger; C099252:oxidized low density lipoprotein; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human