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A Potent Inhibitor of Phosphoinositide 3-Kinase (PI3K) and Mitogen Activated Protein (MAP) Kinase Signalling, Quercetin (3, 3', 4', 5, 7-Pentahydroxyflavone) Promotes Cell Death in Ultraviolet (UV)-B-Irradiated B16F10 Melanoma Cells.

Rafiq RA, Quadri A, Nazir LA, Peerzada K, Ganai BA, Tasduq SA - PLoS ONE (2015)

Bottom Line: The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (ΔΨM).Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-κB nuclear translocation.Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF).

View Article: PubMed Central - PubMed

Affiliation: PK-PD and Toxicology Division, CSIR-Indian Institute of Integrative Medicine, Canal Road, Jammu Tawi, Jammu and Kashmir, India.

ABSTRACT
Ultraviolet (UV) radiation-induced skin damage contributes strongly to the formation of melanoma, a highly lethal form of skin cancer. Quercetin (Qu), the most widely consumed dietary bioflavonoid and well known inhibitor of phosphoinositide 3-kinase (PI3K) and mitogen activated protein (MAP) kinase signalling, has been reported to be chemopreventive in several forms of non-melanoma skin cancers. Here, we report that the treatment of ultraviolet (UV)-B-irradiated B16F10 melanoma cells with quercetin resulted in a dose dependent reduction in cell viability and increased apoptosis. The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (ΔΨM). Promotion of UVB-induced cell death by quercetin was further revealed by cleavage of chromosomal DNA, caspase activation, poly (ADP) ribose polymerase (PARP) cleavage, and an increase in sub-G1 cells. Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-κB nuclear translocation. Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF). Overall, these results suggest the possibility of using quercetin in combination with UVB as a possible treatment option for melanoma in future.

No MeSH data available.


Related in: MedlinePlus

Effect of quercetin on major protein regulators of anti-oxidant defence response in UVB-irradiated B16F10 cells.A, immunoblot analysis of Nrf-2, Catalase and Cu/Zn SOD in B16F10 cells treated with Qu and/or UVB. Signals were quantified for Nrf-2 (B), catalase (C) and Cu/Zn SOD (D) and normalized against β-actin of each band using the Image Lab Software. E, immunoblot analysis of whole cell, cytosolic and nuclear NF-κB in B16F10 cells treated with Qu and/or UVB. F, represents the densitometric analysis of NF-κB (whole cell). G, the signals for cytosolic and nuclear NF-κB were quantified and expressed as ratio of NF-κB(cytosol) / NF-κB(nucleus) for each treatment.
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pone.0131253.g007: Effect of quercetin on major protein regulators of anti-oxidant defence response in UVB-irradiated B16F10 cells.A, immunoblot analysis of Nrf-2, Catalase and Cu/Zn SOD in B16F10 cells treated with Qu and/or UVB. Signals were quantified for Nrf-2 (B), catalase (C) and Cu/Zn SOD (D) and normalized against β-actin of each band using the Image Lab Software. E, immunoblot analysis of whole cell, cytosolic and nuclear NF-κB in B16F10 cells treated with Qu and/or UVB. F, represents the densitometric analysis of NF-κB (whole cell). G, the signals for cytosolic and nuclear NF-κB were quantified and expressed as ratio of NF-κB(cytosol) / NF-κB(nucleus) for each treatment.

Mentions: It has been recognized that exposure of skin to UVB primarily induces the production of superoxide anion radical (O2.-) and hydrogen peroxide (H2O2). Nuclear factor erythroid 2-related factor 2 (Nrf-2) is a key transcription factor in the regulation of antioxidant defence response [11]. We observed that UVB irradiation reduced the expression of Nrf-2. After treatment of UVB–irradiated B16F10 cells with quercetin, we observed a further decrease in the expression of Nrf-2 (Fig 7A and 7B). To elucidate the role of catalase, an enzyme that cleaves H2O2, in response to treatment of UVB–irradiated B16F10 cells with quercetin, we analyzed the expression of catalase by western blotting. It was found that UVB irradiation induced the expression of catalase. In contrast, quercetin decreased the expression of catalase in UVB–irradiated B16F10 cells (Fig 7A and 7C). In addition, UVB irradiation produced an increase in the expression of copper-zinc superoxide dismutase (Cu/Zn SOD), an enzyme that catalyzes the dismutation of superoxide anion radical (O2.-) to dioxygen (O2) and hydrogen peroxide (H2O2). Although the expression of Cu/Zn SOD protein increased upon treatment of UVB–irradiated B16F10 cells with lower doses of quercetin, a gradually decrease was observed at higher doses of quercetin (Fig 7A and 7D). Decrease in Cu/Zn SOD expression following treatment of UVB-irradiated B16F10 cells with higher doses of quercetin is probably due to over consumption of Cu/Zn SOD in response to excessive superoxide anion radical (O2.-) formation. Together, these results indicate that alterations in natural anti-oxidant defence system of cells contribute to oxidative damage and cell death.


A Potent Inhibitor of Phosphoinositide 3-Kinase (PI3K) and Mitogen Activated Protein (MAP) Kinase Signalling, Quercetin (3, 3', 4', 5, 7-Pentahydroxyflavone) Promotes Cell Death in Ultraviolet (UV)-B-Irradiated B16F10 Melanoma Cells.

Rafiq RA, Quadri A, Nazir LA, Peerzada K, Ganai BA, Tasduq SA - PLoS ONE (2015)

Effect of quercetin on major protein regulators of anti-oxidant defence response in UVB-irradiated B16F10 cells.A, immunoblot analysis of Nrf-2, Catalase and Cu/Zn SOD in B16F10 cells treated with Qu and/or UVB. Signals were quantified for Nrf-2 (B), catalase (C) and Cu/Zn SOD (D) and normalized against β-actin of each band using the Image Lab Software. E, immunoblot analysis of whole cell, cytosolic and nuclear NF-κB in B16F10 cells treated with Qu and/or UVB. F, represents the densitometric analysis of NF-κB (whole cell). G, the signals for cytosolic and nuclear NF-κB were quantified and expressed as ratio of NF-κB(cytosol) / NF-κB(nucleus) for each treatment.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4493061&req=5

pone.0131253.g007: Effect of quercetin on major protein regulators of anti-oxidant defence response in UVB-irradiated B16F10 cells.A, immunoblot analysis of Nrf-2, Catalase and Cu/Zn SOD in B16F10 cells treated with Qu and/or UVB. Signals were quantified for Nrf-2 (B), catalase (C) and Cu/Zn SOD (D) and normalized against β-actin of each band using the Image Lab Software. E, immunoblot analysis of whole cell, cytosolic and nuclear NF-κB in B16F10 cells treated with Qu and/or UVB. F, represents the densitometric analysis of NF-κB (whole cell). G, the signals for cytosolic and nuclear NF-κB were quantified and expressed as ratio of NF-κB(cytosol) / NF-κB(nucleus) for each treatment.
Mentions: It has been recognized that exposure of skin to UVB primarily induces the production of superoxide anion radical (O2.-) and hydrogen peroxide (H2O2). Nuclear factor erythroid 2-related factor 2 (Nrf-2) is a key transcription factor in the regulation of antioxidant defence response [11]. We observed that UVB irradiation reduced the expression of Nrf-2. After treatment of UVB–irradiated B16F10 cells with quercetin, we observed a further decrease in the expression of Nrf-2 (Fig 7A and 7B). To elucidate the role of catalase, an enzyme that cleaves H2O2, in response to treatment of UVB–irradiated B16F10 cells with quercetin, we analyzed the expression of catalase by western blotting. It was found that UVB irradiation induced the expression of catalase. In contrast, quercetin decreased the expression of catalase in UVB–irradiated B16F10 cells (Fig 7A and 7C). In addition, UVB irradiation produced an increase in the expression of copper-zinc superoxide dismutase (Cu/Zn SOD), an enzyme that catalyzes the dismutation of superoxide anion radical (O2.-) to dioxygen (O2) and hydrogen peroxide (H2O2). Although the expression of Cu/Zn SOD protein increased upon treatment of UVB–irradiated B16F10 cells with lower doses of quercetin, a gradually decrease was observed at higher doses of quercetin (Fig 7A and 7D). Decrease in Cu/Zn SOD expression following treatment of UVB-irradiated B16F10 cells with higher doses of quercetin is probably due to over consumption of Cu/Zn SOD in response to excessive superoxide anion radical (O2.-) formation. Together, these results indicate that alterations in natural anti-oxidant defence system of cells contribute to oxidative damage and cell death.

Bottom Line: The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (ΔΨM).Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-κB nuclear translocation.Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF).

View Article: PubMed Central - PubMed

Affiliation: PK-PD and Toxicology Division, CSIR-Indian Institute of Integrative Medicine, Canal Road, Jammu Tawi, Jammu and Kashmir, India.

ABSTRACT
Ultraviolet (UV) radiation-induced skin damage contributes strongly to the formation of melanoma, a highly lethal form of skin cancer. Quercetin (Qu), the most widely consumed dietary bioflavonoid and well known inhibitor of phosphoinositide 3-kinase (PI3K) and mitogen activated protein (MAP) kinase signalling, has been reported to be chemopreventive in several forms of non-melanoma skin cancers. Here, we report that the treatment of ultraviolet (UV)-B-irradiated B16F10 melanoma cells with quercetin resulted in a dose dependent reduction in cell viability and increased apoptosis. The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (ΔΨM). Promotion of UVB-induced cell death by quercetin was further revealed by cleavage of chromosomal DNA, caspase activation, poly (ADP) ribose polymerase (PARP) cleavage, and an increase in sub-G1 cells. Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-κB nuclear translocation. Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF). Overall, these results suggest the possibility of using quercetin in combination with UVB as a possible treatment option for melanoma in future.

No MeSH data available.


Related in: MedlinePlus