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Biological responses of three-dimensional cultured fibroblasts by sustained compressive loading include apoptosis and survival activity.

Kanazawa T, Nakagami G, Minematsu T, Yamane T, Huang L, Mugita Y, Noguchi H, Mori T, Sanada H - PLoS ONE (2014)

Bottom Line: Compared to the control, the compressed samples demonstrated that apoptosis was induced in a time- and load- dependent manner; vinculin and stress fiber were scarce; HSP90α, CD44, HAS2, and COX2 expression was upregulated; and the concentrations of HSP90α, hyaluronan (HA), and prostaglandin E2 (PGE2) were increased.In addition, the gene expression of antiapoptotic Bcl2 was significantly increased in the compressed samples compared to the control.These results suggest that compressive loading induces not only apoptosis but also survival activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Gerontological Nursing/Wound Care Management, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

ABSTRACT
Pressure ulcers are characterized by chronicity, which results in delayed wound healing due to pressure. Early intervention for preventing delayed healing due to pressure requires a prediction method. However, no study has reported the prediction of delayed healing due to pressure. Therefore, this study focused on biological response-based molecular markers for the establishment of an assessment technology to predict delayed healing due to pressure. We tested the hypothesis that sustained compressive loading applied to three dimensional cultured fibroblasts leads to upregulation of heat shock proteins (HSPs), CD44, hyaluronan synthase 2 (HAS2), and cyclooxygenase 2 (COX2) along with apoptosis via disruption of adhesion. First, sustained compressive loading was applied to fibroblast-seeded collagen sponges. Following this, collagen sponge samples and culture supernatants were collected for apoptosis and proliferation assays, gene expression analysis, immunocytochemistry, and quantification of secreted substances induced by upregulation of mRNA and protein level. Compared to the control, the compressed samples demonstrated that apoptosis was induced in a time- and load- dependent manner; vinculin and stress fiber were scarce; HSP90α, CD44, HAS2, and COX2 expression was upregulated; and the concentrations of HSP90α, hyaluronan (HA), and prostaglandin E2 (PGE2) were increased. In addition, the gene expression of antiapoptotic Bcl2 was significantly increased in the compressed samples compared to the control. These results suggest that compressive loading induces not only apoptosis but also survival activity. These observations support that HSP90α, HA, and, PGE2 could be potential molecular markers for prediction of delayed wound healing due to pressure.

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The secretions of HSP90α, HA, and PGE2 into cell culture medium were increased by compressive loading.Fibroblasts were seeded to collagen sponge and incubated for 24(A: HSP90α, B: HA, C: PGE2) was measured by ELISA. A value of concentration was normalized by WST-1 value. The results are represented as the mean ± SEM (error bars) of five experiments. Statistical analysis was performed using the Dunnett’s multiple test between non-loaded group and each of loaded group, and statistical significance was taken as p<0.05. A value of p was expressed as: *; p<0.05, **; p<0.01, and ***; p<0.001.
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pone-0104676-g005: The secretions of HSP90α, HA, and PGE2 into cell culture medium were increased by compressive loading.Fibroblasts were seeded to collagen sponge and incubated for 24(A: HSP90α, B: HA, C: PGE2) was measured by ELISA. A value of concentration was normalized by WST-1 value. The results are represented as the mean ± SEM (error bars) of five experiments. Statistical analysis was performed using the Dunnett’s multiple test between non-loaded group and each of loaded group, and statistical significance was taken as p<0.05. A value of p was expressed as: *; p<0.05, **; p<0.01, and ***; p<0.001.

Mentions: We then measured the concentration of HSP90α, HA, and PGE2 in the culture medium. The concentration of HSP90α was significantly higher in the 50, 100, and 200 mmHg groups than in the 0 mmHg group (p = 0.042,  = 0.002, and  = 0.004, respectively; Fig. 5A). The concentration of HA was also significantly higher in the 100 and 200 mmHg groups than in the 0 mmHg group (p = 0.014 and  = 0.021, respectively; Fig. 5B). On the other hand, the concentration of PGE2 was significantly higher only in the 100 mmHg group than in the 0 mmHg group (p = 0.004). An increase in PGE2 was observed in the 50 mmHg and the 200 mmHg groups compared with the 0 mmHg group (p = 0.177 and  = 0.149, respectively; Fig. 5C) but this difference was not statistically significant.


Biological responses of three-dimensional cultured fibroblasts by sustained compressive loading include apoptosis and survival activity.

Kanazawa T, Nakagami G, Minematsu T, Yamane T, Huang L, Mugita Y, Noguchi H, Mori T, Sanada H - PLoS ONE (2014)

The secretions of HSP90α, HA, and PGE2 into cell culture medium were increased by compressive loading.Fibroblasts were seeded to collagen sponge and incubated for 24(A: HSP90α, B: HA, C: PGE2) was measured by ELISA. A value of concentration was normalized by WST-1 value. The results are represented as the mean ± SEM (error bars) of five experiments. Statistical analysis was performed using the Dunnett’s multiple test between non-loaded group and each of loaded group, and statistical significance was taken as p<0.05. A value of p was expressed as: *; p<0.05, **; p<0.01, and ***; p<0.001.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4125229&req=5

pone-0104676-g005: The secretions of HSP90α, HA, and PGE2 into cell culture medium were increased by compressive loading.Fibroblasts were seeded to collagen sponge and incubated for 24(A: HSP90α, B: HA, C: PGE2) was measured by ELISA. A value of concentration was normalized by WST-1 value. The results are represented as the mean ± SEM (error bars) of five experiments. Statistical analysis was performed using the Dunnett’s multiple test between non-loaded group and each of loaded group, and statistical significance was taken as p<0.05. A value of p was expressed as: *; p<0.05, **; p<0.01, and ***; p<0.001.
Mentions: We then measured the concentration of HSP90α, HA, and PGE2 in the culture medium. The concentration of HSP90α was significantly higher in the 50, 100, and 200 mmHg groups than in the 0 mmHg group (p = 0.042,  = 0.002, and  = 0.004, respectively; Fig. 5A). The concentration of HA was also significantly higher in the 100 and 200 mmHg groups than in the 0 mmHg group (p = 0.014 and  = 0.021, respectively; Fig. 5B). On the other hand, the concentration of PGE2 was significantly higher only in the 100 mmHg group than in the 0 mmHg group (p = 0.004). An increase in PGE2 was observed in the 50 mmHg and the 200 mmHg groups compared with the 0 mmHg group (p = 0.177 and  = 0.149, respectively; Fig. 5C) but this difference was not statistically significant.

Bottom Line: Compared to the control, the compressed samples demonstrated that apoptosis was induced in a time- and load- dependent manner; vinculin and stress fiber were scarce; HSP90α, CD44, HAS2, and COX2 expression was upregulated; and the concentrations of HSP90α, hyaluronan (HA), and prostaglandin E2 (PGE2) were increased.In addition, the gene expression of antiapoptotic Bcl2 was significantly increased in the compressed samples compared to the control.These results suggest that compressive loading induces not only apoptosis but also survival activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Gerontological Nursing/Wound Care Management, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

ABSTRACT
Pressure ulcers are characterized by chronicity, which results in delayed wound healing due to pressure. Early intervention for preventing delayed healing due to pressure requires a prediction method. However, no study has reported the prediction of delayed healing due to pressure. Therefore, this study focused on biological response-based molecular markers for the establishment of an assessment technology to predict delayed healing due to pressure. We tested the hypothesis that sustained compressive loading applied to three dimensional cultured fibroblasts leads to upregulation of heat shock proteins (HSPs), CD44, hyaluronan synthase 2 (HAS2), and cyclooxygenase 2 (COX2) along with apoptosis via disruption of adhesion. First, sustained compressive loading was applied to fibroblast-seeded collagen sponges. Following this, collagen sponge samples and culture supernatants were collected for apoptosis and proliferation assays, gene expression analysis, immunocytochemistry, and quantification of secreted substances induced by upregulation of mRNA and protein level. Compared to the control, the compressed samples demonstrated that apoptosis was induced in a time- and load- dependent manner; vinculin and stress fiber were scarce; HSP90α, CD44, HAS2, and COX2 expression was upregulated; and the concentrations of HSP90α, hyaluronan (HA), and prostaglandin E2 (PGE2) were increased. In addition, the gene expression of antiapoptotic Bcl2 was significantly increased in the compressed samples compared to the control. These results suggest that compressive loading induces not only apoptosis but also survival activity. These observations support that HSP90α, HA, and, PGE2 could be potential molecular markers for prediction of delayed wound healing due to pressure.

Show MeSH
Related in: MedlinePlus