ABSTRACT
Title
Molecular mechanisms underlying the anticancer effects of Citrus bergamia juice
Authors
M. R. Ursino1, E. Trapasso1, G. Lo Surdo1, G. Calapai2, M. Navarra1,3
1Dept. Pharmaco-Biological, School of Pharmacy, University of Messina, Italy
2Dept. of Clinical and Experimental Medicine and Pharmacology, School of Medicine, University of Messina, Messina
3IRCCS Centro Neurolesi "Bonino-Pulejo", Messina
1Dept. Pharmaco-Biological, School of Pharmacy, University of Messina, Italy
2Dept. of Clinical and Experimental Medicine and Pharmacology, School of Medicine, University of Messina, Messina
3IRCCS Centro Neurolesi "Bonino-Pulejo", Messina
Abstract
Cancer is a major public health problem worldwide. Both nervous system and liver tumors are two solid malignancies that usually undergo rapid progression with a poor prognosis. Although over the past decade, considerable progress has been made in the multimodality treatment (surgery, chemotherapy, radio-metabolic cure) of both neuroblastoma (NB) and hepatocellular carcinoma (HCC), the healing strategies are still far from satisfaction [1,2]. Thus, development of novel chemotherapeutic and chemopreventive drugs for NB and HCC is important in order to reduce the impact of these diseases.
About 70% of anticancer drugs are natural or semisynthetic products [3], and a lot of evidences underline that natural substances commonly assumed by the diet can have an important role in the prevention of cancer [4].
Citrus Bergamia Risso and Poiteau (Bergamot) is a small tree belonging to the family of Rutaceae, cultivated almost exclusively along the southern coast of Calabria region (Italy). Bergamot fruit is used mostly for the extraction of its essential oil widely used in perfume industry. Bergamot juice (BJ), instead, is considered just a secondary and discarded product. In the light of the growing interest concerning the antiproliferative activities of several fruit juices and vegetable extracts, the object of the present study was to evaluate the effects of BJ on both SH-SY5Y human neuroblastoma and HepG2 human hepatocellular carcinoma cells.
Incubation of HepG2 cells with increased dilution of BJ ranging from 0.5% and 10% for 24, 48 and 72 hours, reduced cell proliferation in time and concentration-dependent manner, as assessed by MTT test. These data were confirmed by the cell growth curves analysis. The reduction of HepG2 growth rate seems to be correlated, at least in part, to a cytotoxic action elicited by BJ, as suggested by the trypan blue assay that show a significant cell death induced by BJ 10%. Moreover, Annexin V staining suggests that BJ is able to activate the programmed cell death, and shows that apoptotic population increases in relationship with the time incubation and BJ concentration. Western blot analysis indicate that apoptosis of HepG2 cells induced by BJ could be due to a decreased expression of Bcl2 and BclXL. Furthermore, BJ increase the expression of P53 that may contribute to the pro-apoptotic activity of BJ and may be responsible for the HepG2 cell cycle arrest in G2 phase.
The same experimental approach was used to assess if the BJ was able to reduce cell proliferation of human neuroblastoma SH-SY5Y. Results of these experiments showed that the antiproliferative effect induced by BJ on this cell line is poorly correlated with increased cell death, but rather it seems to be mediated by a time-and concentration-dependent reduction of cell population in G2 phase. Furthermore, exposure of SH-SY5Y to BJ determined a modification in cellular morphology, causing a marked decrease of cell substrate adhesion.
Our study show that BJ reduces the growth rate of tumor cells from different origin, through the activation of different signaling pathways and molecular mechanisms. In human hepatocellular carcinoma HepG2 cells the antiproliferative activity of BJ is mainly due to a cytotoxic-like effect, while in human neuroblastoma SH-SY5Y cells seems to prevail a cytostatic effect accompanied by a marked reduction of cell adhesion, suggesting a promising role for BJ in the fight against cancer.
[1] Maris (2010) Recent advances in neuroblastoma. The New England Journal of Medicine 362 (23), 2202-2211.
[2] Sanyal et al. (2010) The etiology of hepatocellular carcinoma and consequences for treatment. The Oncologist 15, Suppl 4, 14-22.
[3] Newman and Cragg (2007) Natural products as sources of new drugs over the last 25 years. Journal of Natural Products 70 (3), 461-477.
[4] Amin et al. (2009) Perspectives for cancer prevention with natural compounds. Journal of Clinical Oncology 27 (16), 2712-2725.
About 70% of anticancer drugs are natural or semisynthetic products [3], and a lot of evidences underline that natural substances commonly assumed by the diet can have an important role in the prevention of cancer [4].
Citrus Bergamia Risso and Poiteau (Bergamot) is a small tree belonging to the family of Rutaceae, cultivated almost exclusively along the southern coast of Calabria region (Italy). Bergamot fruit is used mostly for the extraction of its essential oil widely used in perfume industry. Bergamot juice (BJ), instead, is considered just a secondary and discarded product. In the light of the growing interest concerning the antiproliferative activities of several fruit juices and vegetable extracts, the object of the present study was to evaluate the effects of BJ on both SH-SY5Y human neuroblastoma and HepG2 human hepatocellular carcinoma cells.
Incubation of HepG2 cells with increased dilution of BJ ranging from 0.5% and 10% for 24, 48 and 72 hours, reduced cell proliferation in time and concentration-dependent manner, as assessed by MTT test. These data were confirmed by the cell growth curves analysis. The reduction of HepG2 growth rate seems to be correlated, at least in part, to a cytotoxic action elicited by BJ, as suggested by the trypan blue assay that show a significant cell death induced by BJ 10%. Moreover, Annexin V staining suggests that BJ is able to activate the programmed cell death, and shows that apoptotic population increases in relationship with the time incubation and BJ concentration. Western blot analysis indicate that apoptosis of HepG2 cells induced by BJ could be due to a decreased expression of Bcl2 and BclXL. Furthermore, BJ increase the expression of P53 that may contribute to the pro-apoptotic activity of BJ and may be responsible for the HepG2 cell cycle arrest in G2 phase.
The same experimental approach was used to assess if the BJ was able to reduce cell proliferation of human neuroblastoma SH-SY5Y. Results of these experiments showed that the antiproliferative effect induced by BJ on this cell line is poorly correlated with increased cell death, but rather it seems to be mediated by a time-and concentration-dependent reduction of cell population in G2 phase. Furthermore, exposure of SH-SY5Y to BJ determined a modification in cellular morphology, causing a marked decrease of cell substrate adhesion.
Our study show that BJ reduces the growth rate of tumor cells from different origin, through the activation of different signaling pathways and molecular mechanisms. In human hepatocellular carcinoma HepG2 cells the antiproliferative activity of BJ is mainly due to a cytotoxic-like effect, while in human neuroblastoma SH-SY5Y cells seems to prevail a cytostatic effect accompanied by a marked reduction of cell adhesion, suggesting a promising role for BJ in the fight against cancer.
[1] Maris (2010) Recent advances in neuroblastoma. The New England Journal of Medicine 362 (23), 2202-2211.
[2] Sanyal et al. (2010) The etiology of hepatocellular carcinoma and consequences for treatment. The Oncologist 15, Suppl 4, 14-22.
[3] Newman and Cragg (2007) Natural products as sources of new drugs over the last 25 years. Journal of Natural Products 70 (3), 461-477.
[4] Amin et al. (2009) Perspectives for cancer prevention with natural compounds. Journal of Clinical Oncology 27 (16), 2712-2725.