Transforming growth factor-beta induces senescence in hepatocellular carcinoma cells and inhibits tumor growth.
Senescence induction could be used as an effective treatment for hepatocellular carcinoma (HCC). However, major senescence inducers (p53 and p16(Ink4a)) are frequently inactivated in these cancers. We tested whether transforming growth factor-beta (TGF-beta) could serve as a potential senescence inducer in HCC. First, we screened for HCC cell lines with intact TGF-beta signaling that leads to small mothers against decapentaplegic (Smad)-targeted gene activation. Five cell lines met this condition, and all of them displayed a strong senescence response to TGF-beta1 (1-5 ng/mL) treatment. Upon treatment, c-myc was down-regulated, p21(Cip1) and p15(Ink4b) were up-regulated, and cells were arrested at G(1). The expression of p16(Ink4a) was not induced, and the senescence response was independent of p53 status. A short exposure of less than 1 minute was sufficient for a robust senescence response. Forced expression of p21(Cip1) and p15(Ink4b) recapitulated TGF-beta1 effects. Senescence response was associated with reduced nicotinamide adenine dinucleotide phosphate oxidase 4 (Nox4) induction and intracellular reactive oxygen species (ROS) accumulation. The treatment of cells with the ROS scavenger N-acetyl-L-cysteine, or silencing of the NOX4 gene, rescued p21(Cip1) and p15(Ink4b) accumulation as well as the growth arrest in response to TGF-beta. Human HCC tumors raised in immunodeficient mice also displayed TGF-beta1-induced senescence. More importantly, peritumoral injection of TGF-beta1 (2 ng) at 4-day intervals reduced tumor growth by more than 75%. In contrast, the deletion of TGF-beta receptor 2 abolished in vitro senescence response and greatly accelerated in vivo tumor growth.


CONCLUSIONS
TGF-beta induces p53-independent and p16(Ink4a)-independent, but Nox4-dependent, p21(Cip1)-dependent, p15(Ink4b)-dependent, and ROS-dependent senescence arrest in well-differentiated HCC cells. Moreover, TGF-beta-induced senescence in vivo is associated with a strong antitumor response against HCC.
Senturk
Serif
S
BilGen Research Center and Department of Molecular Biology and Genetics, Bilkent University, Ankara, Turkey.
Mumcuoglu
Mine
M
Gursoy-Yuzugullu
Ozge
O
Cingoz
Burcu
B
Akcali
Kamil Can
KC
Ozturk
Mehmet
M
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D006528:Carcinoma, Hepatocellular / Q000503:physiopathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D016922:Cellular Senescence / Q000502:physiology*; D050762:Cyclin-Dependent Kinase Inhibitor p15 / Q000502:physiology; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000502:physiology; D006801:Humans; D008099:Liver / Q000378:metabolism; D008103:Liver Cirrhosis / Q000378:metabolism; D008113:Liver Neoplasms / Q000503:physiopathology*; D051379:Mice; D008819:Mice, Nude; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000502:physiology; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000502:physiology*; D014183:Transplantation, Heterologous
C493956:CDKN1A protein, human; C493966:CDKN2B protein, human; D050762:Cyclin-Dependent Kinase Inhibitor p15; D050759:Cyclin-Dependent Kinase Inhibitor p21; D017382:Reactive Oxygen Species; D016212:Transforming Growth Factor beta; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human