Int J Oral Biol.  2015 Mar;40(1):51-61. 10.11620/IJOB.2015.40.1.051.

Mechanism Underlying Shikonin-induced Apoptosis and Cell Cycle Arrest on SCC25 Human Tongue Squamous Cell Carcinoma Cell Line

• Oh SH
• Park SJ
• Yu SB
• Kim YH
• Kim IR
• Park BS

• KMID: 1789369
• DOI: http://doi.org/10.11620/IJOB.2015.40.1.051

Abstract

Shikonin, a major ingredient in the traditional Chinese herb Lithospermumerythrorhizon, exhibits multiple biological functions including antimicrobial, anti-inflammatory, and antitumor effects. It has recently been reported that shikonin displays antitumor properties in many cancers. This study was aimed to investigate whether shikonin could inhibit oral squamous carcinoma cell (OSCC) growth via mechanisms of apoptosis and cell cycle arrest. The effects of shikonin on the viability and growth of OSCC cell line, SCC25 cells were assessed by MTT assay and clonogenic assays, respectively. Hoechst staining and DNA electrophoresis indicated that the shikonin-treated SCC25 cells were undergoing apoptosis. Western blotting, immunocytochemistry, confocal microscopy, flow cytometry, MMP activity, and proteasome activity also supported the finding that shikonin induces apoptosis. Shikonin treatment of SCC25 cells resulted in a time- and dose-dependent decrease in cell viability, inhibition of cell growth, and increase in apoptotic cell death. The treated SCC25 cells showed several lines of apoptotic manifestation as follows: nuclear condensation; DNA fragmentation; reduced MMP and proteasome activity; decrease in DNA contents; release of cytochrome c into cytosol; translocation of AIF and DFF40 (CAD) onto the nuclei; a significant shift in Bax/Bcl-2 ratio; and activation of caspase-9, -7, -6, and -3, as well as PARP, lamin A/C, and DFF45 (ICAD). Shikonin treatment also resulted in down-regulation of the G1 cell cycle-related proteins and up-regulation of p27(KIP1). Taken together, our present findings demonstrate that shikonin strongly inhibits cell proliferation by modulating the expression of the G1 cell cycle-related proteins, and that it induces apoptosis via the proteasome, mitochondria, and caspase cascades in SCC25 cells.

Keyword

shikonin; apoptosis; human squamous cell carcinoma

MeSH Terms

Apoptosis*
Asian Continental Ancestry Group
Blotting, Western
Carcinoma, Squamous Cell*
Caspase 9
Cell Cycle Checkpoints*
Cell Death
Cell Line*
Cell Proliferation
Cell Survival
Cytochromes c
Cytosol
DNA
DNA Fragmentation
Down-Regulation
Electrophoresis
Flow Cytometry
Humans
Immunohistochemistry
Microscopy, Confocal
Mitochondria
Proteasome Endopeptidase Complex
Tongue*
Up-Regulation
Caspase 9
Cytochromes c
DNA
Proteasome Endopeptidase Complex