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Antitumor and apoptotic effects of quercetin on human melanoma cells involving JNK/P38 MAPK signaling activation
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Introduction
Melanoma is the most malignant type of skin cancer caused by UV rays from the sun. It is easily metastasized to other organs. It accounts for ~4% of all skin cancers, but its mortality rate is as high as 80%. Although melanoma may be treated by surgery, radiation therapy and chemotherapy, the treatments are often ineffective. Despite recent improvements in immunotherapy and targeted therapy, it is difficult to treat because of the side effects caused by drug resistance and high toxicity (Menaa, 2013; White et al., 2009; Megahed et al., 2002). Therefore, over the past several decades, researchers have investigated potential new anticancer substances derived from natural products obtained from plants (Srivastava and Singh, 2004; Ma et al., 2014; Gao et al., 2011).
Quercetin is commonly found in nature, in a variety of foods such as vegetables, tea, fruit and wine Ishizawa et al., 2011). It is also known to have anti-oxidant (Park et al., 2006; Han et al., 2009), anti-inflammatory (Lee et al., 1981; Parveen et al., 2007), antibiosis (Kataoka et al., 2001) as well as anticancer effects, causing apoptosis of prostate cancer cells (Lee et al., 2008a; Vijayababu et al., 2005), liver cancer cells (Tanigawa et al., 2008; Granado-Serrano et al., 2006), colorectal cancer cells (Lim et al., 2007), breast cancer cells (Choi et al., 2008), pancreatic cancer cells (Lee et al., 2013) and lung cancer cells (Lee et al., 2008b). However, the anti-cancer efficacy of quercetin in A375SM, A375P melanoma cells and the underlying molecular mechanisms remain unknown.
Apoptosis is a potential treatment for cancer. It is a type of defense mechanism that prevents the development and progression of cancer by spontaneously removing unnecessary or malfunctioning cells that have DNA damage (Han et al., 2008). Apoptosis inhibits the survival and proliferation of cancer cells and is caused by the Bcl-2 family, which is composed of a pro-apoptotic protein and an anti-apoptotic protein. The Bcl-2 family of proteins play a key role in regulating apoptosis. (Burz et al., 2009).
The mitogen-activated protein kinase cascade (MAPK), which is a mechanism to transmit the extracellular signals to the nucleus in the cell, plays an important role in cell proliferation and differentiation, or apoptosis and survival. MAPK can be classified into extracellular signal-regulated protein kinase (ERK1/2), p38 MAPK, and c-Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK). In general, p38 and JNK are activated in response to extracellular stress and play an important role in the death of inflammatory cells (Yang et al., 2007). In contrast, ERK1/2 activation plays a role in cell proliferation and differentiation but may have anti-apoptotic or pro-apoptotic effects, depending on the nature of the cell (Ballif and Blenis, 2001; Moos and Fitzpatrick, 1998).
Our study investigated the in vitro effect of quercetin on the viability and proliferation of A375SM and A375P melanoma cells, the induction of apoptosis and its mechanism, and quercetin's in vivo effect on inhibiting the proliferation of tumors.
Section snippets
Cell lines and materials
The human melanoma cell lines (A375SM and A375P) for this experiment were purchased from the Korean Cell Line Bank (KCLB, Seoul, Korea). MEM and DMEM were purchased from Welgene (Gyeonsan, Korea). Penicillin-streptomycin, trypsin-EDTA and fetal bovine serum (FBS) were purchased from Gibco BRL (Grand Island, NY, USA). Anti-rabbit IgG, anti-Bax, anti-Bcl-2, anti-β-actin, anti-PARP, anti-JNK, anti-phospho-JNK, anti-ERK1/2, anti-phospho-ERK1/2, anti-p38 and anti-phospho-p38 were purchased from Cell
The effect of quercetin on the viability of melanoma cells
We conducted the MTT assay with quercetin at concentrations of 0, 20, 40, 60, 80, 100, 150 and 200 μM to confirm the effect of quercetin on the survival of A375SM and A375P melanoma cells. When treated with quercetin at different concentrations for 24 h, the survival rate of A375SM melanoma cells was 109% at 20 μM, 89% at 40 μM, 74% at 60 μM, 61% at 80 μM, 59% at 100 μM, 41% at 150 μM, and 32% at 200 μM. When compared to the control group, there was a significant decrease at 40 μM (Fig. 2A).
Discussion
To examine the inhibitory effects of quercetin on the growth and proliferation of A375SM and A375P melanoma cells, each cell was treated with quercetin at concentrations of 0, 20, 40, 60, 80, 100, 150 and 200 μM for 24 h. The MTT assay results showed that the cell viability of the A375SM melanoma cells was 108, 85, 57, 43, 38, 39 and 37% at quercetin concentrations of 0, 20, 40, 60, 80, 100, 150 and 200 μM, respectively. Cell viability decreased significantly at concentrations of 40 μM and
Conclusion
Quercetin significantly decreased the viability and proliferative activity of A375SM cells in a concentration-dependent manner. However, it had no effect on A375P cells. Quercetin showed a significant increase in the stained cells with condensed chromatin due to apoptosis, and the showed a significant increase in apoptosis cells in concentration-dependent manner. Quercetin increased the expression of Bax, cleaved poly-ADP ribose polymerase and phospho-JNK, phospho-p38 and phospho-ERK1/2 while
Funding
This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF, 2017R1A2B4005516).
Conflicts of interest
The authors declare no conflict of interest.
CRediT authorship contribution statement
Sung-Hyun Kim: Conceptualization, Data curation, Project administration, Formal analysis, Writing - original draft, Writing - review & editing. Eun-Seon Yoo: Conceptualization, Validation, Formal analysis. Joong-Seok Woo: Conceptualization, Validation, Formal analysis. So-Hee Han: Investigation, Visualization. Jae-Han Lee: Investigation, Visualization. Soo-Hyun Jung: Investigation, Software. Hyeong-Jin Kim: Conceptualization, Methodology. Ji-Youn Jung: Supervision, Conceptualization,
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