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Apoptosis Induction in Primary Human Colorectal Cancer Cell Lines and Retarded Tumor Growth in SCID Mice by Sulforaphane.

Chen MJ, Tang WY, Hsu CW, Tsai YT, Wu JF, Lin CW, Cheng YM, Hsu YC - Evid Based Complement Alternat Med (2011)

Bottom Line: In addition, treated cells showed nuclear apoptotic morphology that coincided with an activation of caspase-3, and loss of mitochondrial membrane potential (ΔΨm).Incubations at higher SFN doses (12.5 and 25 μM) resulted in cleavage of procaspase-3 and elevated caspase-2, -3, -8, and -9 activity, suggesting that the induction of apoptosis and the sulforaphane-induced mitosis delay at the lower dose are independently regulated.Daily SFN s.c. injections (400 micromol/kg/d for 3 weeks) in severe combined immunodeficient mice with primary human CRC (CP1 to CP5) s.c. tumors resulted in a decrease of mean tumor weight by 70% compared with vehicle-treated controls.

View Article: PubMed Central - PubMed

Affiliation: Division of Traumatology, Department of Surgery, Chi Mei Medical Center, Tainan, Taiwan.

ABSTRACT
We have investigated the anticancer effects of the dietary isothiocyanate sulforaphane (SFN) on colorectal cancer (CRC), using primary cancer cells lines isolated from five Taiwanese colorectal cancer patients as the model for colorectal cancer. SFN-treated cells accumulated in metaphase (SFN 6.25 μM) and subG1 (SFN 12.5 and 25 μM) as determined by flow cytometry. In addition, treated cells showed nuclear apoptotic morphology that coincided with an activation of caspase-3, and loss of mitochondrial membrane potential (ΔΨm). Incubations at higher SFN doses (12.5 and 25 μM) resulted in cleavage of procaspase-3 and elevated caspase-2, -3, -8, and -9 activity, suggesting that the induction of apoptosis and the sulforaphane-induced mitosis delay at the lower dose are independently regulated. Daily SFN s.c. injections (400 micromol/kg/d for 3 weeks) in severe combined immunodeficient mice with primary human CRC (CP1 to CP5) s.c. tumors resulted in a decrease of mean tumor weight by 70% compared with vehicle-treated controls. Our findings suggest that, in addition to the known effects on cancer prevention, sulforaphane may have antitumor activity in established colorectal cancer.

No MeSH data available.


Related in: MedlinePlus

Apoptosis of CRC cell lines induced by SFN. Flow cytometric analysis of PI-Annexin-V to quantify SFN-induced apoptosis in CRC cell lines. (a) Dot plots of five CRC cell lines with SFN treatment at 0, 6.25, 12.5, and 25 μM for 6 h. (b) Results were expressed as a percentage of total apoptosis cells (early and late apoptosis).
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fig3: Apoptosis of CRC cell lines induced by SFN. Flow cytometric analysis of PI-Annexin-V to quantify SFN-induced apoptosis in CRC cell lines. (a) Dot plots of five CRC cell lines with SFN treatment at 0, 6.25, 12.5, and 25 μM for 6 h. (b) Results were expressed as a percentage of total apoptosis cells (early and late apoptosis).

Mentions: Detection between the intact cells, early apoptotic cells, and late apoptotic cells or dead cells could be carried out with PI-annexin-V double staining; thus, we performed this assay to further explore cell apoptosis. To explore the potential role that SFN could play in the apoptosis of CRC cells, the ApopNexin FITC apoptosis detection kit has been used to identify the formation of apoptotic cells in the five CRC cell lines after the 6 hours of exposure to SFN. A typical set of results for the ApopNexin FITC apoptosis detection kit is illustrated in Figure 3(a), in which the annexin V-FITC deposits are indicative of the positive existence of apoptotic cells. A dose-dependent increase in apoptosis was observed (y = 17.62x − 14  R2 = 0.9654), that is, the higher the dose of SFN (6.25, 12.5 and 25 μM) used in the exposure, the greater the extent of apoptosis (Figure 3(b)). The increase of the percentages of apoptotic CRC cell lines was observed in all the doses after treatment for 6 h. In 6 h, approximately 4.6 + 0.6% of five CRC cells were totally apoptotic (early apoptosis and late apoptosis) cells in control. The rate of apoptotic CRC cells increased to 22.9 + 6.5% with 6.25 μM SFN treatments. When the concentrations of SFN increased to 12.5 and 25 μM, the percentages of total apoptotic CRC cells increased to 32.6 + 8.9% and 60.1 + 21.3%, respectively. Taken together, the observations imply that the apoptosis of CRC cell lines is significantly elevated by SFN.


Apoptosis Induction in Primary Human Colorectal Cancer Cell Lines and Retarded Tumor Growth in SCID Mice by Sulforaphane.

Chen MJ, Tang WY, Hsu CW, Tsai YT, Wu JF, Lin CW, Cheng YM, Hsu YC - Evid Based Complement Alternat Med (2011)

Apoptosis of CRC cell lines induced by SFN. Flow cytometric analysis of PI-Annexin-V to quantify SFN-induced apoptosis in CRC cell lines. (a) Dot plots of five CRC cell lines with SFN treatment at 0, 6.25, 12.5, and 25 μM for 6 h. (b) Results were expressed as a percentage of total apoptosis cells (early and late apoptosis).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig3: Apoptosis of CRC cell lines induced by SFN. Flow cytometric analysis of PI-Annexin-V to quantify SFN-induced apoptosis in CRC cell lines. (a) Dot plots of five CRC cell lines with SFN treatment at 0, 6.25, 12.5, and 25 μM for 6 h. (b) Results were expressed as a percentage of total apoptosis cells (early and late apoptosis).
Mentions: Detection between the intact cells, early apoptotic cells, and late apoptotic cells or dead cells could be carried out with PI-annexin-V double staining; thus, we performed this assay to further explore cell apoptosis. To explore the potential role that SFN could play in the apoptosis of CRC cells, the ApopNexin FITC apoptosis detection kit has been used to identify the formation of apoptotic cells in the five CRC cell lines after the 6 hours of exposure to SFN. A typical set of results for the ApopNexin FITC apoptosis detection kit is illustrated in Figure 3(a), in which the annexin V-FITC deposits are indicative of the positive existence of apoptotic cells. A dose-dependent increase in apoptosis was observed (y = 17.62x − 14  R2 = 0.9654), that is, the higher the dose of SFN (6.25, 12.5 and 25 μM) used in the exposure, the greater the extent of apoptosis (Figure 3(b)). The increase of the percentages of apoptotic CRC cell lines was observed in all the doses after treatment for 6 h. In 6 h, approximately 4.6 + 0.6% of five CRC cells were totally apoptotic (early apoptosis and late apoptosis) cells in control. The rate of apoptotic CRC cells increased to 22.9 + 6.5% with 6.25 μM SFN treatments. When the concentrations of SFN increased to 12.5 and 25 μM, the percentages of total apoptotic CRC cells increased to 32.6 + 8.9% and 60.1 + 21.3%, respectively. Taken together, the observations imply that the apoptosis of CRC cell lines is significantly elevated by SFN.

Bottom Line: In addition, treated cells showed nuclear apoptotic morphology that coincided with an activation of caspase-3, and loss of mitochondrial membrane potential (ΔΨm).Incubations at higher SFN doses (12.5 and 25 μM) resulted in cleavage of procaspase-3 and elevated caspase-2, -3, -8, and -9 activity, suggesting that the induction of apoptosis and the sulforaphane-induced mitosis delay at the lower dose are independently regulated.Daily SFN s.c. injections (400 micromol/kg/d for 3 weeks) in severe combined immunodeficient mice with primary human CRC (CP1 to CP5) s.c. tumors resulted in a decrease of mean tumor weight by 70% compared with vehicle-treated controls.

View Article: PubMed Central - PubMed

Affiliation: Division of Traumatology, Department of Surgery, Chi Mei Medical Center, Tainan, Taiwan.

ABSTRACT
We have investigated the anticancer effects of the dietary isothiocyanate sulforaphane (SFN) on colorectal cancer (CRC), using primary cancer cells lines isolated from five Taiwanese colorectal cancer patients as the model for colorectal cancer. SFN-treated cells accumulated in metaphase (SFN 6.25 μM) and subG1 (SFN 12.5 and 25 μM) as determined by flow cytometry. In addition, treated cells showed nuclear apoptotic morphology that coincided with an activation of caspase-3, and loss of mitochondrial membrane potential (ΔΨm). Incubations at higher SFN doses (12.5 and 25 μM) resulted in cleavage of procaspase-3 and elevated caspase-2, -3, -8, and -9 activity, suggesting that the induction of apoptosis and the sulforaphane-induced mitosis delay at the lower dose are independently regulated. Daily SFN s.c. injections (400 micromol/kg/d for 3 weeks) in severe combined immunodeficient mice with primary human CRC (CP1 to CP5) s.c. tumors resulted in a decrease of mean tumor weight by 70% compared with vehicle-treated controls. Our findings suggest that, in addition to the known effects on cancer prevention, sulforaphane may have antitumor activity in established colorectal cancer.

No MeSH data available.


Related in: MedlinePlus