Effect of protein kinase C alpha, caspase-3, and survivin on apoptosis of oral cancer cells induced by staurosporine.
OBJECTIVE
To elucidate inhibition of protein kinase C alpha (PKC alpha) activity by staurosporine on apoptosis of oral cancer cell line tongue squamous cell carcinoma (TSCCa) cells and to clarify the role of survivin and caspase-3 in mediating apoptosis.


METHODS
TSCCa cell viability was measured by MTT assay after 100 nmol/L staurosporine treatment. Apoptotic cells were identified by using phase contrast microscopy, acridine orange/ethidium bromide staining, and flow cytometry. Level of PKC alpha and its subcellular location were investigated using Western blot analysis. Expression of survivin and caspase-3 were evaluated using immunocytochemistry.


RESULTS
Staurosporine significantly inhibited the cell viability of TSCCa cells in a dose- and time-dependent manner. Marked cell accumulation in G2/M phase was observed after 100 nmol/L staurosporine exposure for 6 h and 12 h. In addition, the percentage of apoptosis increased in a time-dependent manner, from 2.9% in control cultures to approximately 27.4% at 100 nmol/L staurosporine treatment for 24 h. Staurosporine displayed difference in inhibitory efficacy between cytosolic and membrance-derived PKC alpha. The content of PKCalpha in membrane versus cytosol decreased quickly, from 0.45 in ethanol-treated control cultures to 0.18 after staurosporine exposure for 24 h (P<0.01). After treatment with staurosporine, a time-dependent reduction of survivin and an activation of caspase-3 were observed in TSCCa cells.


CONCLUSIONS
Staurosporine inhibited cell viability and promoted apoptosis in TSCCa cells. Inhibition of PKCalpha activity might be a potential mechanism for staurosporine to induce apoptosis in this cell line. The cleavage of survivin and activation of caspase-3 signaling pathway might contribute to PKC alpha inhibition-induced apoptosis.
Zhang
Yu-xia
YX
Department of Pathology and Pathophysiology, School of Medicine, Wuhan University, Wuhan 430071, China.
Yu
Shi-bin
SB
Ou-Yang
Jing-ping
JP
Xia
Dong
D
Wang
Min
M
Li
Jin-rong
JR
D017209:Apoptosis / Q000187:drug effects; D002294:Carcinoma, Squamous Cell / Q000378:metabolism / Q000473:pathology; D053148:Caspase 3 / Q000378:metabolism*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000494:pharmacology; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism*; D051571:Protein Kinase C-alpha / Q000378:metabolism*; D019311:Staurosporine / Q000008:administration & dosage / Q000494:pharmacology*; D000077022:Survivin; D014062:Tongue Neoplasms / Q000378:metabolism* / Q000473:pathology
C107489:BIRC5 protein, human; D004791:Enzyme Inhibitors; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D000077022:Survivin; D051571:Protein Kinase C-alpha; D053148:Caspase 3; D019311:Staurosporine