shikonin and thiazolyl-blue

The aim of this study was to examine the anti-invasion effect of Shikonin on human high-metastatic adenoid cystic carcinoma (ACC-M) cells and to explain the possible molecular mechanism involved.. The ACC-M cells were treated with Shikonin (0, 2.5, 5, 10 μM) for 24 h. The protein levels and gelatinolytic activities of MMP-2 and MMP-9 were analyzed using Western blot and Gelatin zymography test, respectively. Matrigel invasion assays were used to investigate tumor invasive potential and electromobility shift assays were used to determine the activity of NF-κB.. The invasiveness of ACC-M cells was reduced in a dose dependent manner following 24-h treatment of up to 10 μM of the Shikonin at which concentration no cytotoxicity occurred. The protein levels and gelatinolytic activities of MMP-9 were significantly suppressed by increasing Shikonin concentrations. The down-regulation of MMP-9 appeared to be via the inactivation of NF-κB as the treatment with Shikonin suppressed the protein level of phosphate-IkBa, which was accompanied by a decrease in DNA-binding level of the factor.. Shikonin inhibits tumor invasion via downregulation of MMP-9 expression in ACC-M cells. Pharmacologic inhibition of the NF-κB-mediated MMP-9 expression by Shikonin might be a powerful treatment option for ACC patients in future.

Topics: Antineoplastic Agents; Blotting, Western; Carcinoma, Adenoid Cystic; Cell Culture Techniques; Cell Line, Tumor; Cell Survival; Coloring Agents; Dose-Response Relationship, Drug; Down-Regulation; Drugs, Chinese Herbal; Electrophoresis, Polyacrylamide Gel; Humans; I-kappa B Proteins; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Naphthoquinones; Neoplasm Invasiveness; NF-kappa B; Tetrazolium Salts; Thiazoles; Time Factors; Tumor Necrosis Factor-alpha

Beta,beta-dimethyl acryl shikonin is an extract from the root of plant Arnebia nobilis which has been shown to possess anti-cancer activity. However, its toxicity limited further development of shikonin as a therapeutic agent. Subsequently, several analogues of beta,beta-dimethyl acryl shikonin were synthesized. One of these analogues, shikonin 93/637 was found to be significantly less toxic compared to shikonin. This study is aimed to determine the cell cycle associated differences in the susceptibility of U937 cells to apoptosis induced by shikonin analogue 93/637 (SA). Lower concentrations of SA (approximately 100 nM) showed no significant changes in cell growth. However, higher concentrations (approximately 500 nM) resulted in growth inhibition of U937 cells after 48 h of treatment with SA as measured by MTT assay. Flow cytometric analysis showed that SA treatment resulted in blocking of cell cycle progression in G1 phase. Decreased expression of Cyclin D, CDK 4 and PCNA was observed with SA treatment corroborating the G1 block. DNA gel electrophoresis showed an oligonucleotide ladder pattern, a distinct characteristic of DNA fragmentation associated with programmed cell death. Ribonuclease protection assay revealed inhibition of bcl2 expression at transcriptional level. SA treatment also resulted in induction of caspase-3 activity. The results suggest the involvement of bcl2 and Caspase-3 in SA induced apoptosis of human U937 cells.

Topics: Apoptosis; Caspase 3; Cell Survival; Cinnamates; DNA Fragmentation; Enzyme Activation; Flow Cytometry; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Naphthoquinones; Neoplasms; Ribonucleases; Tetrazolium Salts; Thiazoles; Transcription, Genetic; U937 Cells