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Clinical significance of stromal apoptosis in colorectal cancer.

Koelink PJ, Sier CF, Hommes DW, Lamers CB, Verspaget HW - Br. J. Cancer (2009)

Bottom Line: Epithelial and stromal apoptosis, as well as total apoptosis, were significantly higher in CRC compared with corresponding adjacent normal mucosa.Epithelial apoptosis was not associated with clinical outcome.In contrast, low stromal apoptosis (< or = median caspase-3/M30) was found to be an independent prognostic factor for overall survival, disease-free survival and disease recurrence, with HRs (95% CI) of 1.66 (1.17-2.35), 1.62 (1.15-2.29) and 1.69 (1.01-2.85), respectively.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology-Hepatology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.

ABSTRACT

Background: Epithelial and stromal cells play an important role in the development of colorectal cancer (CRC). We aimed to determine the prognostic significance of both epithelial and stromal cell apoptosis in CRC.

Methods: Total apoptosis was determined by caspase-3 activity measurements in protein homogenates of CRC specimens and adjacent normal mucosa of 211 CRC patients. Epithelial apoptosis was determined by an ELISA specific for a caspase-3-degraded cytokeratin 18 product, the M30 antigen. Stromal apoptosis was determined from the ratio between total and epithelial apoptosis.

Results: Epithelial and stromal apoptosis, as well as total apoptosis, were significantly higher in CRC compared with corresponding adjacent normal mucosa. Low total tumour apoptosis (< or = median caspase-3 activity) was associated with a significantly worse disease recurrence (hazard ratio (HR), 95% confidence interval (95% CI): 1.77 (1.05-3.01)), independent of clinocopathological parameters. Epithelial apoptosis was not associated with clinical outcome. In contrast, low stromal apoptosis (< or = median caspase-3/M30) was found to be an independent prognostic factor for overall survival, disease-free survival and disease recurrence, with HRs (95% CI) of 1.66 (1.17-2.35), 1.62 (1.15-2.29) and 1.69 (1.01-2.85), respectively.

Interpretation: Stromal apoptosis, in contrast to epithelial apoptosis, is an important factor with respect to survival and disease-recurrence in CRC.

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Caspase-3 activity in CRC. CRC specimens with high caspase-3 activity show levels of the active form of caspase-3, whereas low caspase-3 activity CRC specimens only show inactive (pro-) caspase-3 as determined by immunoblotting for total caspase-3 (A). Caspase-3 activity and M30 antigen in CRC show a weak but significant correlation (B). Some tumours show high caspase-3 activity without high levels of M30 antigen, suggesting stromal apoptosis. Photomicrographs of immunohistochemical stainings for active caspase-3, M30 and pan-cytokeratin showing tumours with stromal cells expressing active caspase-3 (C), as represented by Tumour 1 (left panel), and tumours with mainly epithelial cells expressing active caspase-3, correlating with M30 staining (represented by Tumour 2, right panel). Scale bars=100 μm.
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fig1: Caspase-3 activity in CRC. CRC specimens with high caspase-3 activity show levels of the active form of caspase-3, whereas low caspase-3 activity CRC specimens only show inactive (pro-) caspase-3 as determined by immunoblotting for total caspase-3 (A). Caspase-3 activity and M30 antigen in CRC show a weak but significant correlation (B). Some tumours show high caspase-3 activity without high levels of M30 antigen, suggesting stromal apoptosis. Photomicrographs of immunohistochemical stainings for active caspase-3, M30 and pan-cytokeratin showing tumours with stromal cells expressing active caspase-3 (C), as represented by Tumour 1 (left panel), and tumours with mainly epithelial cells expressing active caspase-3, correlating with M30 staining (represented by Tumour 2, right panel). Scale bars=100 μm.

Mentions: Immunoblot analysis confirmed that tumours with a high caspase-3 activity had high protein levels of active caspase-3 compared with low caspase-3 activity tumours (Figure 1A). Overall there was a weak but significant correlation between tumour caspase-3 activity and tumour M30 antigen levels, as expected, because the M30 antigen is generated by active caspase-3 (Figure 1B). Some tumours had really high caspase-3 activity levels, without high M30 antigen levels, indicating high stromal apoptosis. Immunohistochemical analysis confirmed that in these tumours active caspase-3 was indeed mainly expressed by diverse non-epithelial cells and were almost M30 antigen-negative (Figure 1C, represented by Tumour 1), compared with other tumours in which epithelial cells were expressing most of the active caspase-3 and M30 antigen-positive (represented by tumour 2). Some high power magnifications of tumour 1 to illustrate the diverse nature of stromal apoptosis are shown in Supplementary Figure 1. Our measurements enabled us to further differentiate in cell type origin of apoptosis by calculating the percentage of apoptotic epithelial cells through the formula M30/CK18 × 100 and for stromal apoptosis through the formula caspase-3/M30. This epithelial as well as stromal apoptosis was also significantly higher in tumour tissue compared with corresponding normal tissue (Table 1).


Clinical significance of stromal apoptosis in colorectal cancer.

Koelink PJ, Sier CF, Hommes DW, Lamers CB, Verspaget HW - Br. J. Cancer (2009)

Caspase-3 activity in CRC. CRC specimens with high caspase-3 activity show levels of the active form of caspase-3, whereas low caspase-3 activity CRC specimens only show inactive (pro-) caspase-3 as determined by immunoblotting for total caspase-3 (A). Caspase-3 activity and M30 antigen in CRC show a weak but significant correlation (B). Some tumours show high caspase-3 activity without high levels of M30 antigen, suggesting stromal apoptosis. Photomicrographs of immunohistochemical stainings for active caspase-3, M30 and pan-cytokeratin showing tumours with stromal cells expressing active caspase-3 (C), as represented by Tumour 1 (left panel), and tumours with mainly epithelial cells expressing active caspase-3, correlating with M30 staining (represented by Tumour 2, right panel). Scale bars=100 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2736838&req=5

fig1: Caspase-3 activity in CRC. CRC specimens with high caspase-3 activity show levels of the active form of caspase-3, whereas low caspase-3 activity CRC specimens only show inactive (pro-) caspase-3 as determined by immunoblotting for total caspase-3 (A). Caspase-3 activity and M30 antigen in CRC show a weak but significant correlation (B). Some tumours show high caspase-3 activity without high levels of M30 antigen, suggesting stromal apoptosis. Photomicrographs of immunohistochemical stainings for active caspase-3, M30 and pan-cytokeratin showing tumours with stromal cells expressing active caspase-3 (C), as represented by Tumour 1 (left panel), and tumours with mainly epithelial cells expressing active caspase-3, correlating with M30 staining (represented by Tumour 2, right panel). Scale bars=100 μm.
Mentions: Immunoblot analysis confirmed that tumours with a high caspase-3 activity had high protein levels of active caspase-3 compared with low caspase-3 activity tumours (Figure 1A). Overall there was a weak but significant correlation between tumour caspase-3 activity and tumour M30 antigen levels, as expected, because the M30 antigen is generated by active caspase-3 (Figure 1B). Some tumours had really high caspase-3 activity levels, without high M30 antigen levels, indicating high stromal apoptosis. Immunohistochemical analysis confirmed that in these tumours active caspase-3 was indeed mainly expressed by diverse non-epithelial cells and were almost M30 antigen-negative (Figure 1C, represented by Tumour 1), compared with other tumours in which epithelial cells were expressing most of the active caspase-3 and M30 antigen-positive (represented by tumour 2). Some high power magnifications of tumour 1 to illustrate the diverse nature of stromal apoptosis are shown in Supplementary Figure 1. Our measurements enabled us to further differentiate in cell type origin of apoptosis by calculating the percentage of apoptotic epithelial cells through the formula M30/CK18 × 100 and for stromal apoptosis through the formula caspase-3/M30. This epithelial as well as stromal apoptosis was also significantly higher in tumour tissue compared with corresponding normal tissue (Table 1).

Bottom Line: Epithelial and stromal apoptosis, as well as total apoptosis, were significantly higher in CRC compared with corresponding adjacent normal mucosa.Epithelial apoptosis was not associated with clinical outcome.In contrast, low stromal apoptosis (< or = median caspase-3/M30) was found to be an independent prognostic factor for overall survival, disease-free survival and disease recurrence, with HRs (95% CI) of 1.66 (1.17-2.35), 1.62 (1.15-2.29) and 1.69 (1.01-2.85), respectively.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology-Hepatology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.

ABSTRACT

Background: Epithelial and stromal cells play an important role in the development of colorectal cancer (CRC). We aimed to determine the prognostic significance of both epithelial and stromal cell apoptosis in CRC.

Methods: Total apoptosis was determined by caspase-3 activity measurements in protein homogenates of CRC specimens and adjacent normal mucosa of 211 CRC patients. Epithelial apoptosis was determined by an ELISA specific for a caspase-3-degraded cytokeratin 18 product, the M30 antigen. Stromal apoptosis was determined from the ratio between total and epithelial apoptosis.

Results: Epithelial and stromal apoptosis, as well as total apoptosis, were significantly higher in CRC compared with corresponding adjacent normal mucosa. Low total tumour apoptosis (< or = median caspase-3 activity) was associated with a significantly worse disease recurrence (hazard ratio (HR), 95% confidence interval (95% CI): 1.77 (1.05-3.01)), independent of clinocopathological parameters. Epithelial apoptosis was not associated with clinical outcome. In contrast, low stromal apoptosis (< or = median caspase-3/M30) was found to be an independent prognostic factor for overall survival, disease-free survival and disease recurrence, with HRs (95% CI) of 1.66 (1.17-2.35), 1.62 (1.15-2.29) and 1.69 (1.01-2.85), respectively.

Interpretation: Stromal apoptosis, in contrast to epithelial apoptosis, is an important factor with respect to survival and disease-recurrence in CRC.

Show MeSH
Related in: MedlinePlus