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The leukemia inhibitory factor (LIF) and p21 mediate the TGFβ tumor suppressive effects in human cutaneous melanoma.

Humbert L, Ghozlan M, Canaff L, Tian J, Lebrun JJ - BMC Cancer (2015)

Bottom Line: Techniques involved immunoblotting, immunohistochemistry, real time PCR and luciferase reporter assays.Interestingly, we also showed that TGFβ-mediated LIF expression is required for TGFβ-induced cell cycle arrest and caspase-mediated apoptosis, as well as for TGFβ-mediated inhibition of cell migration.Moreover, we found that TGFβ-mediated LIF expression leads to activation of transcription of the cell cycle inhibitor p21 in a STAT3-dependent manner, and further showed that p21 is required for TGFβ/LIF-mediated cell cycle arrest and TGFβ-induced gene activation of several pro-apoptotic genes.

View Article: PubMed Central - PubMed

Affiliation: Division of Medical Oncology, Department of Medicine, McGill University Health Centre, Montreal, QC, Canada. laure.humbert@mail.mcgill.ca.

ABSTRACT

Background: Cutaneous melanoma is the most lethal skin cancer and its incidence in developed countries has dramatically increased over the past decades. Localized tumors are easily treated by surgery, but advanced melanomas lack efficient treatment and are associated with very poor outcomes. Thus, understanding the processes underlying melanoma development and progression is critical. The Transforming Growth Factor beta (TGFβ) acts as a potent tumor suppressor in human melanoma, by inhibiting cell growth and preventing cellular migration and invasion.

Methods: In this study, we aimed at elucidating the molecular mechanisms underlying TGFβ-mediated tumor suppression. Human cutaneous melanoma cell lines, derived from different patients, were used to assess for cell cycle analysis, apoptosis/caspase activity and cell migration. Techniques involved immunoblotting, immunohistochemistry, real time PCR and luciferase reporter assays.

Results: We found the leukemia inhibitory factor (LIF) to be strongly up-regulated by TGFβ in melanoma cells, defining LIF as a novel TGFβ downstream target gene in cutaneous melanoma. Interestingly, we also showed that TGFβ-mediated LIF expression is required for TGFβ-induced cell cycle arrest and caspase-mediated apoptosis, as well as for TGFβ-mediated inhibition of cell migration. Moreover, we found that TGFβ-mediated LIF expression leads to activation of transcription of the cell cycle inhibitor p21 in a STAT3-dependent manner, and further showed that p21 is required for TGFβ/LIF-mediated cell cycle arrest and TGFβ-induced gene activation of several pro-apoptotic genes.

Conclusions: Together, our results define the LIF/p21 signaling cascade as a novel tumor suppressive-like pathway in melanoma, acting downstream of TGFβ to regulate cell cycle arrest and cell death, further highlight new potential therapeutic strategies for the treatment of cutaneous melanoma.

No MeSH data available.


Related in: MedlinePlus

The TGFβ/LIF pathway is lost across melanoma progression. Tissue sections from a melanoma microarray slide were stained for P-Smad3 and LIF levels. The staining was scored from 0 to 4: (0, no staining of neoplastic cells, 1, weak staining, 2, weak to moderate staining, 3, moderate to strong staining, and 4, strong staining). Representative pictures were taken at the magnification 40X (upper panel). Data is graphed as the percentage of samples with levels 0-2 or 3-4 for each category (lower panel).
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Fig6: The TGFβ/LIF pathway is lost across melanoma progression. Tissue sections from a melanoma microarray slide were stained for P-Smad3 and LIF levels. The staining was scored from 0 to 4: (0, no staining of neoplastic cells, 1, weak staining, 2, weak to moderate staining, 3, moderate to strong staining, and 4, strong staining). Representative pictures were taken at the magnification 40X (upper panel). Data is graphed as the percentage of samples with levels 0-2 or 3-4 for each category (lower panel).

Mentions: Having highlighted the TGFβ/LIF pathway as a potent tumor suppressor in melanoma, we then assessed the clinical relevance of these findings. For this, we analyzed phospho-Smad3 and LIF expression levels by immunohistochemistry in a tissue microarray consisting of 24 benign tumors, 56 malignant tumors, and 20 metastatic melanomas. Phospho-Smad3 was measured as an indicator of TGFβ signaling activity in these tumors. As shown in Figure 6, neoplastic cells in benign tumors showed high levels of phosphorylated Smad3, indicative of high TGFβ signaling activities. However, phospho-Smad3 levels were reduced in malignant tumors and even further diminished in the metastatic tissue samples. This is consistent with previous findings showing that normal melanocytes in culture are sensitive to the growth inhibitory effects of TGFβ, whereas melanoma cell lines demonstrate various degrees of resistance to TGFβ inhibitory effects, proportional to the tumor progression stage [14,15]. Interestingly, LIF expression levels exhibited a similar pattern, showing very high levels of expression in benign tumors, with a progressive decrease in malignant and metastatic tumors. These results indicate that while the TGFβ/LIF signaling axis acts as tumor suppressor pathway in the early stages of melanoma progression, it is partially disrupted in advanced stages, further emphasizing its essential role in the prevention of melanoma development and progression.Figure 6


The leukemia inhibitory factor (LIF) and p21 mediate the TGFβ tumor suppressive effects in human cutaneous melanoma.

Humbert L, Ghozlan M, Canaff L, Tian J, Lebrun JJ - BMC Cancer (2015)

The TGFβ/LIF pathway is lost across melanoma progression. Tissue sections from a melanoma microarray slide were stained for P-Smad3 and LIF levels. The staining was scored from 0 to 4: (0, no staining of neoplastic cells, 1, weak staining, 2, weak to moderate staining, 3, moderate to strong staining, and 4, strong staining). Representative pictures were taken at the magnification 40X (upper panel). Data is graphed as the percentage of samples with levels 0-2 or 3-4 for each category (lower panel).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4389797&req=5

Fig6: The TGFβ/LIF pathway is lost across melanoma progression. Tissue sections from a melanoma microarray slide were stained for P-Smad3 and LIF levels. The staining was scored from 0 to 4: (0, no staining of neoplastic cells, 1, weak staining, 2, weak to moderate staining, 3, moderate to strong staining, and 4, strong staining). Representative pictures were taken at the magnification 40X (upper panel). Data is graphed as the percentage of samples with levels 0-2 or 3-4 for each category (lower panel).
Mentions: Having highlighted the TGFβ/LIF pathway as a potent tumor suppressor in melanoma, we then assessed the clinical relevance of these findings. For this, we analyzed phospho-Smad3 and LIF expression levels by immunohistochemistry in a tissue microarray consisting of 24 benign tumors, 56 malignant tumors, and 20 metastatic melanomas. Phospho-Smad3 was measured as an indicator of TGFβ signaling activity in these tumors. As shown in Figure 6, neoplastic cells in benign tumors showed high levels of phosphorylated Smad3, indicative of high TGFβ signaling activities. However, phospho-Smad3 levels were reduced in malignant tumors and even further diminished in the metastatic tissue samples. This is consistent with previous findings showing that normal melanocytes in culture are sensitive to the growth inhibitory effects of TGFβ, whereas melanoma cell lines demonstrate various degrees of resistance to TGFβ inhibitory effects, proportional to the tumor progression stage [14,15]. Interestingly, LIF expression levels exhibited a similar pattern, showing very high levels of expression in benign tumors, with a progressive decrease in malignant and metastatic tumors. These results indicate that while the TGFβ/LIF signaling axis acts as tumor suppressor pathway in the early stages of melanoma progression, it is partially disrupted in advanced stages, further emphasizing its essential role in the prevention of melanoma development and progression.Figure 6

Bottom Line: Techniques involved immunoblotting, immunohistochemistry, real time PCR and luciferase reporter assays.Interestingly, we also showed that TGFβ-mediated LIF expression is required for TGFβ-induced cell cycle arrest and caspase-mediated apoptosis, as well as for TGFβ-mediated inhibition of cell migration.Moreover, we found that TGFβ-mediated LIF expression leads to activation of transcription of the cell cycle inhibitor p21 in a STAT3-dependent manner, and further showed that p21 is required for TGFβ/LIF-mediated cell cycle arrest and TGFβ-induced gene activation of several pro-apoptotic genes.

View Article: PubMed Central - PubMed

Affiliation: Division of Medical Oncology, Department of Medicine, McGill University Health Centre, Montreal, QC, Canada. laure.humbert@mail.mcgill.ca.

ABSTRACT

Background: Cutaneous melanoma is the most lethal skin cancer and its incidence in developed countries has dramatically increased over the past decades. Localized tumors are easily treated by surgery, but advanced melanomas lack efficient treatment and are associated with very poor outcomes. Thus, understanding the processes underlying melanoma development and progression is critical. The Transforming Growth Factor beta (TGFβ) acts as a potent tumor suppressor in human melanoma, by inhibiting cell growth and preventing cellular migration and invasion.

Methods: In this study, we aimed at elucidating the molecular mechanisms underlying TGFβ-mediated tumor suppression. Human cutaneous melanoma cell lines, derived from different patients, were used to assess for cell cycle analysis, apoptosis/caspase activity and cell migration. Techniques involved immunoblotting, immunohistochemistry, real time PCR and luciferase reporter assays.

Results: We found the leukemia inhibitory factor (LIF) to be strongly up-regulated by TGFβ in melanoma cells, defining LIF as a novel TGFβ downstream target gene in cutaneous melanoma. Interestingly, we also showed that TGFβ-mediated LIF expression is required for TGFβ-induced cell cycle arrest and caspase-mediated apoptosis, as well as for TGFβ-mediated inhibition of cell migration. Moreover, we found that TGFβ-mediated LIF expression leads to activation of transcription of the cell cycle inhibitor p21 in a STAT3-dependent manner, and further showed that p21 is required for TGFβ/LIF-mediated cell cycle arrest and TGFβ-induced gene activation of several pro-apoptotic genes.

Conclusions: Together, our results define the LIF/p21 signaling cascade as a novel tumor suppressive-like pathway in melanoma, acting downstream of TGFβ to regulate cell cycle arrest and cell death, further highlight new potential therapeutic strategies for the treatment of cutaneous melanoma.

No MeSH data available.


Related in: MedlinePlus