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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 attenuates PI3K-Akt pathway and MAPK signalling in UVB-irradiated B16F10 melanoma cells.A, western blot analysis of B-raf, p-MEK, MEK1/2, p-ERK, ERK1/2, phospho-p38, p-38, phospho-JNK and JNK in cells treated with Qu and/or UVB. β-actin was used as loading control. B, C, D, E and F represent the densitometric analysis of B-raf, p-MEK/MEK, p-ERK/ERK, phospho-p38/p-38, phospho-JNK/JNK respectively. G, immunoblot analysis of PI3K-α and p-Akt in B16F10 cells at 24 h post-UVB and/ or Qu treatment. Signals were quantified for PI3K (H) and p-Akt (I) using Image Lab Software (Bio Rad). *, 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.g008: Quercetin attenuates PI3K-Akt pathway and MAPK signalling in UVB-irradiated B16F10 melanoma cells.A, western blot analysis of B-raf, p-MEK, MEK1/2, p-ERK, ERK1/2, phospho-p38, p-38, phospho-JNK and JNK in cells treated with Qu and/or UVB. β-actin was used as loading control. B, C, D, E and F represent the densitometric analysis of B-raf, p-MEK/MEK, p-ERK/ERK, phospho-p38/p-38, phospho-JNK/JNK respectively. G, immunoblot analysis of PI3K-α and p-Akt in B16F10 cells at 24 h post-UVB and/ or Qu treatment. Signals were quantified for PI3K (H) and p-Akt (I) using Image Lab Software (Bio Rad). *, 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: The mitogen-activated protein (MAP) kinase pathway, virtually activated in all melanomas, regulates cell proliferation, metastasis and survival [43]. We studied the effect of quercetin on MAPK signalling in UVB-irradiated B16F10 melanoma cells through western blotting. UVB irradiation of B16F10 cells decreased the expression of B-raf, phospho-MEK and, phospho-ERK. Treatment of UVB-irradiated B16F10 cells with quercetin induced a further decrease in the expression of phospho-MEK and its downstream effector phospho-ERK (Fig 8A–8D). Interestingly, phosphorylation of stress kinase p-38 increased upon treatment of UVB–irradiated B16F10 cells with quercetin (Fig 8A and 8E). However, quercetin caused no significant increase in UVB–induced phosphorylation of JNK (Fig 8A and 8F). Furthermore, treatment of UVB-irradiated B16F10 cells with quercetin markedly decreased the expression of PI3K and phosphorylation of its downstream effector Akt (Fig 8G–8I). Overall, these results suggest that combined UVB and quercetin treatment modulates mitogen-activated protein (MAP) kinase family proteins and attenuates PI3K/Akt survival signals in favour of 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 attenuates PI3K-Akt pathway and MAPK signalling in UVB-irradiated B16F10 melanoma cells.A, western blot analysis of B-raf, p-MEK, MEK1/2, p-ERK, ERK1/2, phospho-p38, p-38, phospho-JNK and JNK in cells treated with Qu and/or UVB. β-actin was used as loading control. B, C, D, E and F represent the densitometric analysis of B-raf, p-MEK/MEK, p-ERK/ERK, phospho-p38/p-38, phospho-JNK/JNK respectively. G, immunoblot analysis of PI3K-α and p-Akt in B16F10 cells at 24 h post-UVB and/ or Qu treatment. Signals were quantified for PI3K (H) and p-Akt (I) using Image Lab Software (Bio Rad). *, 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.g008: Quercetin attenuates PI3K-Akt pathway and MAPK signalling in UVB-irradiated B16F10 melanoma cells.A, western blot analysis of B-raf, p-MEK, MEK1/2, p-ERK, ERK1/2, phospho-p38, p-38, phospho-JNK and JNK in cells treated with Qu and/or UVB. β-actin was used as loading control. B, C, D, E and F represent the densitometric analysis of B-raf, p-MEK/MEK, p-ERK/ERK, phospho-p38/p-38, phospho-JNK/JNK respectively. G, immunoblot analysis of PI3K-α and p-Akt in B16F10 cells at 24 h post-UVB and/ or Qu treatment. Signals were quantified for PI3K (H) and p-Akt (I) using Image Lab Software (Bio Rad). *, 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: The mitogen-activated protein (MAP) kinase pathway, virtually activated in all melanomas, regulates cell proliferation, metastasis and survival [43]. We studied the effect of quercetin on MAPK signalling in UVB-irradiated B16F10 melanoma cells through western blotting. UVB irradiation of B16F10 cells decreased the expression of B-raf, phospho-MEK and, phospho-ERK. Treatment of UVB-irradiated B16F10 cells with quercetin induced a further decrease in the expression of phospho-MEK and its downstream effector phospho-ERK (Fig 8A–8D). Interestingly, phosphorylation of stress kinase p-38 increased upon treatment of UVB–irradiated B16F10 cells with quercetin (Fig 8A and 8E). However, quercetin caused no significant increase in UVB–induced phosphorylation of JNK (Fig 8A and 8F). Furthermore, treatment of UVB-irradiated B16F10 cells with quercetin markedly decreased the expression of PI3K and phosphorylation of its downstream effector Akt (Fig 8G–8I). Overall, these results suggest that combined UVB and quercetin treatment modulates mitogen-activated protein (MAP) kinase family proteins and attenuates PI3K/Akt survival signals in favour of 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