Additive effects of C(2)-ceramide on paclitaxel-induced premature senescence of human lung cancer cells.
OBJECTIVE
the aims of the study are to investigate the additive effect of exogenous short-carbon chain phospholipids, C(2)-ceramide, on an anti-cancer drug paclitaxel (PTX)-induced senescence of human non-small cell lung cancer (NSCLC) cells deficient in functional p53 and p16, and to examine whether mitogen-activated protein kinase (MAPK) plays a role in ceramide-sensitized senescence of NSCLC cells.


METHODS
to determine whether exogenous C(2)-ceramide renders lung cancer cells more sensitive to PTX treatment, techniques employing a flow cytometry-based cell cycle analysis and acidic β-galactosidase staining for senescent cells were used. Furthermore, to elucidate the role of MAPK proteins in modulating senescence, assays for protein levels of selective MAPKs and Bcl-2 family members, and detection of transcriptional levels senescence-associated genes were used in the study.


RESULTS
a sub-lethal dose of C(2)-ceramide sensitized the NSCLC H1299 cells to PTX treatment. The additive effects of C(2)-ceramide and PTX resulted in proliferative inhibition, G(2)-phase arrest of cell cycle, activation of p38 and eventually premature senescence. Importantly, neither p53, p21(waf1/cip1) nor p16(ink4) was shown to be involved in C(2)-ceramide-sensitized proliferative inhibition and senescence of H1299 cells by PTX in our study.


CONCLUSIONS
our study demonstrates that the short-carbon chain C(2)-ceramide can effectively sensitize PTX-induced senescence of H1299 cells via both p21(waf1/cip1)- and p16(ink4)-independent pathways.
Chen
Jeff Yi-Fu
JY
Department of Biotechnology, Center of Excellence for Environmental Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan.
Hwang
Chi-Ching
CC
Chen
Wei-Yi
WY
Lee
Jing-Ching
JC
Fu
Tzu-Fun
TF
Fang
Kang
K
Chu
Ying-Chieh
YC
Huang
Ya-Lan
YL
Lin
Jia-Cheng
JC
Tsai
Wen-Hui
WH
Chang
Hsueh-Wei
HW
Chen
Bing-Hung
BH
Chiu
Chien-Chih
CC
D017209:Apoptosis / Q000187:drug effects; D000075747:Baculoviral IAP Repeat-Containing 3 Protein; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D016922:Cellular Senescence / Q000187:drug effects*; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics; D053938:DNA Fragmentation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D016195:G2 Phase / Q000187:drug effects; D019204:GTP-Binding Proteins / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008840:Microfilament Proteins / Q000235:genetics; D009124:Muscle Proteins / Q000235:genetics; D015676:Osteonectin / Q000235:genetics; D017239:Paclitaxel / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D011503:Transglutaminases / Q000235:genetics; D044767:Ubiquitin-Protein Ligases; D001616:beta-Galactosidase / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D051034:Inhibitor of Apoptosis Proteins; D008840:Microfilament Proteins; D009124:Muscle Proteins; C064769:N-acetylsphingosine; D015676:Osteonectin; D017395:Plasminogen Activator Inhibitor 1; D019253:Proto-Oncogene Proteins c-bcl-2; C051321:SERPINE1 protein, human; C051615:transgelin; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; C494832:BIRC3 protein, human; D000075747:Baculoviral IAP Repeat-Containing 3 Protein; D044767:Ubiquitin-Protein Ligases; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D001616:beta-Galactosidase; D053148:Caspase 3; D019204:GTP-Binding Proteins; D013110:Sphingosine; D017239:Paclitaxel