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Cell-line-specific stimulation of tumor cell aggressiveness by wound healing factors - a central role for STAT3.

Ekblad L, Lindgren G, Persson E, Kjellén E, Wennerberg J - BMC Cancer (2013)

Bottom Line: These effects were accompanied by a sharp increase in activation of signal transducer and activator of transcription 3 (STAT3).The involvement of c-Met or epidermal growth factor receptor family members was excluded, while the interleukin-6 receptor was found to be partly responsible for the activation of STAT3.In conclusion, we found cell-line-specific effects of wound healing factors on HNSCC, setting the stage for therapy development and predictive opportunities.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oncology, Lund University, Lund, Sweden. Lars.Ekblad@med.lu.se

ABSTRACT

Background: Local recurrence is a major factor affecting survival after treatment for head and neck squamous cell carcinoma (HNSCC). It is possible that the normal processes involved in wound healing after surgical removal of a primary tumor can boost the regrowth of residual cancer cells, thereby contributing to the recurrent growth. In this work, we collected human wound fluids and used them to investigate the effect of wound healing factors on HNSCC cell lines in vitro.

Methods: Wound fluids were collected from thyroidectomized patients diagnosed with benign disease and were included in assays of cell proliferation, migration, cell scattering, and invasion. The involvement of intracellular signaling pathways and membrane receptors were investigated by western blotting and the inclusion of specific inhibitors.

Results: One out of four cell lines was greatly stimulated in proliferation, migration, cell scattering, and invasion by the addition of wound fluid as compared with addition of fetal bovine or human serum. These effects were accompanied by a sharp increase in activation of signal transducer and activator of transcription 3 (STAT3). Inhibition of STAT3 activation abolished the wound fluid response, showing that STAT3 plays an important role in the wound healing response. Several of the observed phenotypic changes were epithelial-to-mesenchymal transition (EMT)-like, but the appropriate changes were not seen in any of the EMT markers investigated. The involvement of c-Met or epidermal growth factor receptor family members was excluded, while the interleukin-6 receptor was found to be partly responsible for the activation of STAT3.

Conclusions: In conclusion, we found cell-line-specific effects of wound healing factors on HNSCC, setting the stage for therapy development and predictive opportunities.

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The effect of HWF on cell scattering of HN-7 cells. (a) At time 0 the medium was changed to DMEM containing 10% FBS, HS, or HWF as indicated. The cells were photographed with a 10× objective at the specified time points. (b) Photographs sampled as under A were analyzed in ImageJ software as stated in Materials and Methods to determine the area covered by cell colonies (N=16). (c) The exponential area increase in the later time points in panel B was determined by non-linear regression and subtracted from the total area increase, yielding an approximation of the scatter contribution (N=16). (d) As in panel B, cells were exposed to 10% HWF at time 0, but after approximately 4 h the medium was changed back to 10% FBS (N=8). (e) The cells were exposed to 10% HWF collected at different time points after operation (OP). The area increase during the first hour was determined as in panel B (N=8). Error bars represent SEM.
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Figure 2: The effect of HWF on cell scattering of HN-7 cells. (a) At time 0 the medium was changed to DMEM containing 10% FBS, HS, or HWF as indicated. The cells were photographed with a 10× objective at the specified time points. (b) Photographs sampled as under A were analyzed in ImageJ software as stated in Materials and Methods to determine the area covered by cell colonies (N=16). (c) The exponential area increase in the later time points in panel B was determined by non-linear regression and subtracted from the total area increase, yielding an approximation of the scatter contribution (N=16). (d) As in panel B, cells were exposed to 10% HWF at time 0, but after approximately 4 h the medium was changed back to 10% FBS (N=8). (e) The cells were exposed to 10% HWF collected at different time points after operation (OP). The area increase during the first hour was determined as in panel B (N=8). Error bars represent SEM.

Mentions: In the migration experiments, the HN-7 cells seemed to detach from each other at the migration front when treated with HWF. To investigate this scattering phenomenon further, we seeded cells at low density and changed medium when the cells were approximately 25% confluent. The cells were then photographed at intervals during 24 h. Addition of medium with FBS did not affect the appearance of the cells appreciably. With HS, the area of the colonies seemed to increase somewhat initially (within 3 h) and some cells tended to adopt a more spindle-shaped morphology. These changes reverted after another two hours. The cell colonies that were exposed to HWF expanded visibly within three hours, and several cells detached from the colonies and became spindle-shaped. This effect persisted during the whole observation period (Figure 2a).


Cell-line-specific stimulation of tumor cell aggressiveness by wound healing factors - a central role for STAT3.

Ekblad L, Lindgren G, Persson E, Kjellén E, Wennerberg J - BMC Cancer (2013)

The effect of HWF on cell scattering of HN-7 cells. (a) At time 0 the medium was changed to DMEM containing 10% FBS, HS, or HWF as indicated. The cells were photographed with a 10× objective at the specified time points. (b) Photographs sampled as under A were analyzed in ImageJ software as stated in Materials and Methods to determine the area covered by cell colonies (N=16). (c) The exponential area increase in the later time points in panel B was determined by non-linear regression and subtracted from the total area increase, yielding an approximation of the scatter contribution (N=16). (d) As in panel B, cells were exposed to 10% HWF at time 0, but after approximately 4 h the medium was changed back to 10% FBS (N=8). (e) The cells were exposed to 10% HWF collected at different time points after operation (OP). The area increase during the first hour was determined as in panel B (N=8). Error bars represent SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3585883&req=5

Figure 2: The effect of HWF on cell scattering of HN-7 cells. (a) At time 0 the medium was changed to DMEM containing 10% FBS, HS, or HWF as indicated. The cells were photographed with a 10× objective at the specified time points. (b) Photographs sampled as under A were analyzed in ImageJ software as stated in Materials and Methods to determine the area covered by cell colonies (N=16). (c) The exponential area increase in the later time points in panel B was determined by non-linear regression and subtracted from the total area increase, yielding an approximation of the scatter contribution (N=16). (d) As in panel B, cells were exposed to 10% HWF at time 0, but after approximately 4 h the medium was changed back to 10% FBS (N=8). (e) The cells were exposed to 10% HWF collected at different time points after operation (OP). The area increase during the first hour was determined as in panel B (N=8). Error bars represent SEM.
Mentions: In the migration experiments, the HN-7 cells seemed to detach from each other at the migration front when treated with HWF. To investigate this scattering phenomenon further, we seeded cells at low density and changed medium when the cells were approximately 25% confluent. The cells were then photographed at intervals during 24 h. Addition of medium with FBS did not affect the appearance of the cells appreciably. With HS, the area of the colonies seemed to increase somewhat initially (within 3 h) and some cells tended to adopt a more spindle-shaped morphology. These changes reverted after another two hours. The cell colonies that were exposed to HWF expanded visibly within three hours, and several cells detached from the colonies and became spindle-shaped. This effect persisted during the whole observation period (Figure 2a).

Bottom Line: These effects were accompanied by a sharp increase in activation of signal transducer and activator of transcription 3 (STAT3).The involvement of c-Met or epidermal growth factor receptor family members was excluded, while the interleukin-6 receptor was found to be partly responsible for the activation of STAT3.In conclusion, we found cell-line-specific effects of wound healing factors on HNSCC, setting the stage for therapy development and predictive opportunities.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oncology, Lund University, Lund, Sweden. Lars.Ekblad@med.lu.se

ABSTRACT

Background: Local recurrence is a major factor affecting survival after treatment for head and neck squamous cell carcinoma (HNSCC). It is possible that the normal processes involved in wound healing after surgical removal of a primary tumor can boost the regrowth of residual cancer cells, thereby contributing to the recurrent growth. In this work, we collected human wound fluids and used them to investigate the effect of wound healing factors on HNSCC cell lines in vitro.

Methods: Wound fluids were collected from thyroidectomized patients diagnosed with benign disease and were included in assays of cell proliferation, migration, cell scattering, and invasion. The involvement of intracellular signaling pathways and membrane receptors were investigated by western blotting and the inclusion of specific inhibitors.

Results: One out of four cell lines was greatly stimulated in proliferation, migration, cell scattering, and invasion by the addition of wound fluid as compared with addition of fetal bovine or human serum. These effects were accompanied by a sharp increase in activation of signal transducer and activator of transcription 3 (STAT3). Inhibition of STAT3 activation abolished the wound fluid response, showing that STAT3 plays an important role in the wound healing response. Several of the observed phenotypic changes were epithelial-to-mesenchymal transition (EMT)-like, but the appropriate changes were not seen in any of the EMT markers investigated. The involvement of c-Met or epidermal growth factor receptor family members was excluded, while the interleukin-6 receptor was found to be partly responsible for the activation of STAT3.

Conclusions: In conclusion, we found cell-line-specific effects of wound healing factors on HNSCC, setting the stage for therapy development and predictive opportunities.

Show MeSH
Related in: MedlinePlus