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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

Quercetin promotes UVB-induced mitochondrial membrane potential (ΔΨM) dissipation and modulates the expression of various pro- and anti-apoptotic proteins.A, analysis of mitochondrial membrane potential (ΔΨM) in B16F10 cells treated with Qu and/or UVB. B, represents the fold increase in the percentage of ΔΨm low cells relative to control cells. C, immunoblot analysis of Bax, Bcl-2, Bim and AIF in B16F10 cells treated with Qu and/or UVB. D, signals were quantified for Bax and Bcl-2 and expressed as Bcl-2 (total)/Bax (total) ratio for each treatment. E and F, represents the densitometric analysis of Bim and AIF respectively. *, P<0.05; **, P<0.01 for control versus treatments; #, P<0.05, ##, P<0.01 for control versus UVB-alone treatment versus UVB + Qu treatments.
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pone.0131253.g004: Quercetin promotes UVB-induced mitochondrial membrane potential (ΔΨM) dissipation and modulates the expression of various pro- and anti-apoptotic proteins.A, analysis of mitochondrial membrane potential (ΔΨM) in B16F10 cells treated with Qu and/or UVB. B, represents the fold increase in the percentage of ΔΨm low cells relative to control cells. C, immunoblot analysis of Bax, Bcl-2, Bim and AIF in B16F10 cells treated with Qu and/or UVB. D, signals were quantified for Bax and Bcl-2 and expressed as Bcl-2 (total)/Bax (total) ratio for each treatment. E and F, represents the densitometric analysis of Bim and AIF respectively. *, P<0.05; **, P<0.01 for control versus treatments; #, P<0.05, ##, P<0.01 for control versus UVB-alone treatment versus UVB + Qu treatments.

Mentions: In addition to being the cell’s powerhouse, the site of adenosine triphosphate (ATP) synthesis, mitochondria can also be the source of signals that can initiate programmed cell death [35]. The majority of ATP synthesis in living cells is driven by the electrochemical gradient built across the inner mitochondrial membrane known as mitochondrial membrane potential, symbolized as ΔΨM. During apoptosis, the mitochondrial membrane potential (ΔΨM) of a cell falls, facilitating the release of cytochrome c and activation of downstream cellular apoptotic machinery [36]. Here, we have studied the effect of quercetin on the mitochondrial physiology of UVB-irradiated B16F10 cells. Flow cytometry showed that 5 mJ/cm2 UVB caused 2.5-fold increase in the percentage of ΔΨM low cells relative to control. However, the treatment of UVB-irradiated B16F10 cells with 5, 10, 20 and 40 μM quercetin caused 2.6-, 5-, 5.1- and 7.5-fold increase in the percentage of ΔΨM low cells (Fig 4A and 4B). During UVB irradiation, depolarization of mitochondrial membrane potential (ΔΨM) is regulated by the proteins of Bcl-2 family [12] and the outcome of death signal usually depends on the balance between the positive and negative apoptotic regulators of the Bcl-2 family [37]. UVB irradiation decreased the ratio of Bcl-2 to that of Bax. Interestingly, quercetin caused significant and gradual decrease in the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF) (Fig 4C–4F). These changes in the protein expression of Bcl-2 family proteins and mitochondria dysfunction may account for the increased apoptosis.


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)

Quercetin promotes UVB-induced mitochondrial membrane potential (ΔΨM) dissipation and modulates the expression of various pro- and anti-apoptotic proteins.A, analysis of mitochondrial membrane potential (ΔΨM) in B16F10 cells treated with Qu and/or UVB. B, represents the fold increase in the percentage of ΔΨm low cells relative to control cells. C, immunoblot analysis of Bax, Bcl-2, Bim and AIF in B16F10 cells treated with Qu and/or UVB. D, signals were quantified for Bax and Bcl-2 and expressed as Bcl-2 (total)/Bax (total) ratio for each treatment. E and F, represents the densitometric analysis of Bim and AIF respectively. *, P<0.05; **, P<0.01 for control versus treatments; #, P<0.05, ##, P<0.01 for control versus UVB-alone treatment versus UVB + Qu treatments.
© Copyright Policy
Related In: Results  -  Collection

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pone.0131253.g004: Quercetin promotes UVB-induced mitochondrial membrane potential (ΔΨM) dissipation and modulates the expression of various pro- and anti-apoptotic proteins.A, analysis of mitochondrial membrane potential (ΔΨM) in B16F10 cells treated with Qu and/or UVB. B, represents the fold increase in the percentage of ΔΨm low cells relative to control cells. C, immunoblot analysis of Bax, Bcl-2, Bim and AIF in B16F10 cells treated with Qu and/or UVB. D, signals were quantified for Bax and Bcl-2 and expressed as Bcl-2 (total)/Bax (total) ratio for each treatment. E and F, represents the densitometric analysis of Bim and AIF respectively. *, P<0.05; **, P<0.01 for control versus treatments; #, P<0.05, ##, P<0.01 for control versus UVB-alone treatment versus UVB + Qu treatments.
Mentions: In addition to being the cell’s powerhouse, the site of adenosine triphosphate (ATP) synthesis, mitochondria can also be the source of signals that can initiate programmed cell death [35]. The majority of ATP synthesis in living cells is driven by the electrochemical gradient built across the inner mitochondrial membrane known as mitochondrial membrane potential, symbolized as ΔΨM. During apoptosis, the mitochondrial membrane potential (ΔΨM) of a cell falls, facilitating the release of cytochrome c and activation of downstream cellular apoptotic machinery [36]. Here, we have studied the effect of quercetin on the mitochondrial physiology of UVB-irradiated B16F10 cells. Flow cytometry showed that 5 mJ/cm2 UVB caused 2.5-fold increase in the percentage of ΔΨM low cells relative to control. However, the treatment of UVB-irradiated B16F10 cells with 5, 10, 20 and 40 μM quercetin caused 2.6-, 5-, 5.1- and 7.5-fold increase in the percentage of ΔΨM low cells (Fig 4A and 4B). During UVB irradiation, depolarization of mitochondrial membrane potential (ΔΨM) is regulated by the proteins of Bcl-2 family [12] and the outcome of death signal usually depends on the balance between the positive and negative apoptotic regulators of the Bcl-2 family [37]. UVB irradiation decreased the ratio of Bcl-2 to that of Bax. Interestingly, quercetin caused significant and gradual decrease in the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF) (Fig 4C–4F). These changes in the protein expression of Bcl-2 family proteins and mitochondria dysfunction may account for the increased apoptosis.

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