Nestin serves as a prosurvival determinant that is linked to the cytoprotective effect of epidermal growth factor in rat vascular smooth muscle cells.
Nestin is an intermediate filament protein mainly expressed in muscle and neural progenitors. Recently, we reported that nestin is expressed in rat vascular smooth muscle cells (VSMCs), disappears after serum-deprivation and then is re-expressed again following EGF stimulation. As the function of nestin in VSMCs remains unknown, its anti-apoptotic function was investigated in this study. We first showed that cell viability of nestin-depleted cells following H(2)O(2) treatments decreased by nestin RNAi. Further DNA laddering analysis and flow cytometry results demonstrated that this loss of cell viability was mediated through apoptosis. In addition, caspase-9, caspase-3 and PARP were activated in nestin-depleted VSMCs following H(2)O(2) treatments, indicating that nestin has an upstream inhibitory effect on caspase activation. It is well known that EGF serves as a survival factor in rat VSMCs. Here, we show that the cytoprotective effect of EGF was prevented by nestin RNAi. In addition, the inhibition of Cdk5 prevented Bcl-2 phosphorylation and enhanced H(2)O(2)-induced caspase-3 activation as well as subsequent DNA fragmentation. Taken together, these results provide evidence for another cytoprotective role of EGF in that it is mediated through its stimulation of nestin expression which leads to the prevention of caspase activation by Cdk-5-induced Bcl-2 phosphorylation in rat VSMCs.
Huang
Yuan-Li
YL
Institute of Molecular and Cellular Biology, National Taiwan University, No. 1, Sec. 4, Roosevelt Road, Taipei 106, Taiwan.
Wu
Ching-Ming
CM
Shi
Guey-Yueh
GY
Wu
Georgiana Cho-Chen
GC
Lee
Hsinyu
H
Jiang
Meei-Jyh
MJ
Wu
Hua-Lin
HL
Yang
Hsi-Yuan
HY
D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D051360:Cyclin-Dependent Kinase 5 / Q000037:antagonists & inhibitors / Q000378:metabolism; D019610:Cytoprotection / Q000502:physiology*; D004815:Epidermal Growth Factor / Q000502:physiology*; D005434:Flow Cytometry; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D007381:Intermediate Filament Proteins / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D064231:Nestin; D010766:Phosphorylation; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D034741:RNA, Small Interfering; D051381:Rats
D051017:Apoptosis Regulatory Proteins; D061945:Caspase Inhibitors; D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; C577486:Nes protein, rat; D064231:Nestin; D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering; D004815:Epidermal Growth Factor; D011065:Poly(ADP-ribose) Polymerases; D051360:Cyclin-Dependent Kinase 5; C505410:Casp3 protein, rat; C506435:Casp9 protein, rat; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases