Caffeic Acid Induces Intrinsic Apoptotic Pathway in MG-63 Osteosarcoma Cells Through Bid Truncation and Cytochrome c Release

Abstract

BACKGROUND: Caffeic acid has been reported to induce apoptosis in MG-63 osteosarcoma cells via caspases activation. However, apoptotic pathway that is involved in the caffeic acid-induced apoptosis is still unclear. Present study aimed to investigate the role of cytochrome c (Cyt c) release and BH3-interacting death (Bid) activation in caffeic acid-induced apoptosis in MG-63 osteosarcoma cells.METHODS: MG-63 cells were cultured, pretreated with/without Z-VAD FMK and treated with/without 10 μg/mL caffeic acid. Treated MG-63 cells were then lysed, homogenized, and processed further to prepare cell lysate and mitochondrial fraction. Immunoblotting method was used to measure the amount of Bid and truncated Bid (t-Bid) as well as mitochondrial and cytosolic Cyt c.RESULTS: The amount of Bid and mitochondrial Cyt c in MG-63 cells decreased in a time-dependent manner, while the amount of t-Bid and cytosolic Cyt c increased in a time-dependent manner. By pretreatment of 100 μM Z-VAD-FMK for 2 h, the amount of Bid and mitochondrial Cyt c was significantly higher, while the amount of t-Bid and cytosolic Cyt c was significantly lower after caffeic acid treatment for 6 and 12 h compared to MG-63 cells that were not pretreated.CONCLUSION: Caffeic acid could induce Cyt c release through the activation of Bid in MG-63 osteosarcoma cells.KEYWORDS: caffeic acid, osteosarcoma, MG-63 cells, Bid, t-Bid, cytochrome c, Z-VAD-FMK

Caffeic Acid Inhibits Tumour Mass Formation in MG-63 Cells-induced Nude Mice

“…This result suggested that caffeic acid might induces MG-63 cells into apoptosis. (3,11) Only few studies regarding this was conducted on MG-63 cells-induced mice, however the result of this current study is in line with a study on B7-P815 cellsinduced mice which also showed that ingested caffeic acid significantly hinders tumour growth in the terms of size. (20) Previous study on multiple myeloma cells, ARH-77, also shows that a derivative of caffeic acid, namely the caffeic acid phenethyl ester, is able to inhibit the tumour growth and induces cell death in a dose-and time-dependent manner.…”

“…This result suggested that caffeic acid pre-treatment promote the cell death of MG-63 population. (3) Bigger mass nodule in the subcutaneous area was also observed in the untreated MG-63 cells-induced mice compared to the one that received intra-tumour injection of caffeic acid. This proved that the effectiveness of caffeic acid mL caffeic acid, even though not significant, yet obvious inhibition of tumour nodule formation was observed, which was shown by a smaller visible mass (with average of 12.71 mm 3 ) (Figure 2B).…”

“…After 90 days, to observe the formation of visible tumour mass, the mice were anaesthetised, and the picture of the subcutaneous area was taken for observation and measurement. The mass nodule volume was measured according to previous study (17) to obtain the tumour mass volume presented in mm 3 . In each group, the experiments were repeated 3 times.…”

“…After 6 times of caffeic acid injection, mice were anaesthetised, and the picture of visible tumour mass were taken for observation and measurement. The tumour mass nodule volume was also measured according to previous study (17) and presented in mm 3 . In each group, the experiments were repeated 3 times.…”

Caffeic Acid Induces Apoptosis in MG-63 Osteosarcoma Cells via Protein Kinase C Delta (PKCδ) Translocation and Mitochondrial Membrane Potential Reduction

“…(19,20) Studies have shown that caffeic acid triggers MG-63 osteosarcoma cell apoptosis by activating caspases (21,22) and may affect intrinsic and extrinsic apoptotic pathways. (23) Protein kinase C delta (PKCδ) has been reported to be involved in apoptosis of several cancer cells, including lung (24) and prostate cancer. (25) This protein is translocated from cytosol to various organelles, including mitochondria, in the presence of apoptotic stimuli, such as anticancer agents.…”