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Sanguisorba officinalis L synergistically enhanced 5-fluorouracil cytotoxicity in colorectal cancer cells by promoting a reactive oxygen species-mediated, mitochondria-caspase-dependent apoptotic pathway

View Article: PubMed Central - PubMed

ABSTRACT

Sanguisorba officinalis L. radix is a widely used herb called DiYu (DY) in China and has an extensive range of bioactivities, including anti-cancer, anti-inflammatory, and anti-oxidative activities. However, there is little evidence to support its anti-cancer effects against colorectal cancer (CRC). The first-line chemotherapeutic agent 5-fluorouracil (5-FU) is used to treat CRC, but its efficiency is hampered by acquired drug resistance. This study found that a water extract of DY exerted anti-proliferative effects against two CRC cell lines (HCT-116 and RKO), and it sensitized CRC cells to 5-FU therapy by activating a reactive oxygen species (ROS)-mediated, mitochondria-caspase-dependent apoptotic pathway. Co-treatment of DY and 5-FU significantly elevated ROS levels, up-regulated Bax/Bcl-2 ratio and triggered mitochondrial dysfunction, followed by a release of cytochrome c and up-regulation of proteins such as cleaved-caspase-9/3 and cleaved-PARP. Additionally, the induction of autophagy may be involved in mediating synergism of DY in HCT-116 cells. Gallic acid (GA), catechinic acid (CA) and ellagic acid (EA) were identified as the potential chief constituents responsible for the synergistic effects of DY. In conclusion, co-treatment of DY, specifically GA, CA and EA, with 5-FU may be a potential alternative therapeutic strategy for CRC by enhancing an intrinsic apoptotic pathway.

No MeSH data available.


Three tannins in DY dramatically sensitized CRC cells to the 5-FU therapy.(a) Cells were treated with GA, CA or EA separately or in a combination, cell viabilities were evaluate by CCK8 assay. *P < 0.05, **P < 0.01, ***P < 0.001, compared with (GA + CA + EA) groups. (b) Concentration-effect curves of three tannins and DY were plotted, *P < 0.05, **P < 0.01, ***P < 0.001, vs DY groups. (c) The synergistic anti-proliferative effects of the three tannins and DY on the cytotoxicity of 5-FU in CRC cells. #P < 0.05, ##P < 0.01, ###P < 0.001, vs (DY + 5-FU) group. While *P < 0.05, **P < 0.01, ***P < 0.001, vs (GA + CA + EA + 5-FU) group. All the data are expressed as mean ± SD (n = 3). (d) CI and DRI values from different combined groups are demonstrated in the heat-maps plotted by R programming, which are assessed by the color bars on the right (→red, values are greater; →blue, values are lower; 5-FU1 and 5-FU2 represent DRIs in the DY + 5-FU and tanins+5-FU groups, respectively; F30–F90, 30% to 90% effective rate).
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f6: Three tannins in DY dramatically sensitized CRC cells to the 5-FU therapy.(a) Cells were treated with GA, CA or EA separately or in a combination, cell viabilities were evaluate by CCK8 assay. *P < 0.05, **P < 0.01, ***P < 0.001, compared with (GA + CA + EA) groups. (b) Concentration-effect curves of three tannins and DY were plotted, *P < 0.05, **P < 0.01, ***P < 0.001, vs DY groups. (c) The synergistic anti-proliferative effects of the three tannins and DY on the cytotoxicity of 5-FU in CRC cells. #P < 0.05, ##P < 0.01, ###P < 0.001, vs (DY + 5-FU) group. While *P < 0.05, **P < 0.01, ***P < 0.001, vs (GA + CA + EA + 5-FU) group. All the data are expressed as mean ± SD (n = 3). (d) CI and DRI values from different combined groups are demonstrated in the heat-maps plotted by R programming, which are assessed by the color bars on the right (→red, values are greater; →blue, values are lower; 5-FU1 and 5-FU2 represent DRIs in the DY + 5-FU and tanins+5-FU groups, respectively; F30–F90, 30% to 90% effective rate).

Mentions: Tannins are the major components in DY (~17% weight of radix)21. Gallic acid (GA) and ellagic acid (EA) are representative hydrolysable tannins, while catechinic acid is one of the oligomeric procyanidins in DY. In this study, a water extract of DY was prepared using an extraction method with good reproducibility (RSD% of GA content = 1.39%, Supplementary Fig. 1a), and the three tannins in the overall extract of DY were separated and identified by HPLC analysis (Fig. 5 and Supplementary Fig. 1b). As determined by HPLC, the three tannins accounted for 3.9% of the total content of the DY extract. To investigate whether GA, CA and EA were the major bioactive constituents of DY that were responsible for the anti-proliferative effects on the CRC cell lines, we evaluated the cytotoxicity of each tannin in CRC cells. No obvious inhibitory activity was observed for any of the tannins (IC50 > 100 μM). Then, we concurrently treated the cells with two or three tannins at same concentrations in 100 μg/ml DY extracts. The cell viabilities were significantly inhibited by the combined tannins (P < 0.05, vs single tannin group), and treatments with two or three tannins showed comparable potency in inhibiting the CRC cells (we chose the combination of three tannins for further research considering its better cooperation). However, the inhibitory effects were lower than that of the overall DY extract (P < 0.05) (Fig. 6a). The dose-effect curves of the three tannins are shown in Fig. 6b. The HCT-116 cells were more sensitive to the tannin treatment than the RKO cells, the growth of which was notably inhibited in a dose-dependent manner. The inhibitory effects of the tannins and the DY extract were comparable at low concentrations (25–50 μg/ml). However, with increased concentrations, the anti-proliferative effects of the tannins turned to be much lower than that of the DY treatment. These results indicate that GA, CA and EA together could effectively inhibit the CRC cells but can’t replace DY.


Sanguisorba officinalis L synergistically enhanced 5-fluorouracil cytotoxicity in colorectal cancer cells by promoting a reactive oxygen species-mediated, mitochondria-caspase-dependent apoptotic pathway
Three tannins in DY dramatically sensitized CRC cells to the 5-FU therapy.(a) Cells were treated with GA, CA or EA separately or in a combination, cell viabilities were evaluate by CCK8 assay. *P < 0.05, **P < 0.01, ***P < 0.001, compared with (GA + CA + EA) groups. (b) Concentration-effect curves of three tannins and DY were plotted, *P < 0.05, **P < 0.01, ***P < 0.001, vs DY groups. (c) The synergistic anti-proliferative effects of the three tannins and DY on the cytotoxicity of 5-FU in CRC cells. #P < 0.05, ##P < 0.01, ###P < 0.001, vs (DY + 5-FU) group. While *P < 0.05, **P < 0.01, ***P < 0.001, vs (GA + CA + EA + 5-FU) group. All the data are expressed as mean ± SD (n = 3). (d) CI and DRI values from different combined groups are demonstrated in the heat-maps plotted by R programming, which are assessed by the color bars on the right (→red, values are greater; →blue, values are lower; 5-FU1 and 5-FU2 represent DRIs in the DY + 5-FU and tanins+5-FU groups, respectively; F30–F90, 30% to 90% effective rate).
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f6: Three tannins in DY dramatically sensitized CRC cells to the 5-FU therapy.(a) Cells were treated with GA, CA or EA separately or in a combination, cell viabilities were evaluate by CCK8 assay. *P < 0.05, **P < 0.01, ***P < 0.001, compared with (GA + CA + EA) groups. (b) Concentration-effect curves of three tannins and DY were plotted, *P < 0.05, **P < 0.01, ***P < 0.001, vs DY groups. (c) The synergistic anti-proliferative effects of the three tannins and DY on the cytotoxicity of 5-FU in CRC cells. #P < 0.05, ##P < 0.01, ###P < 0.001, vs (DY + 5-FU) group. While *P < 0.05, **P < 0.01, ***P < 0.001, vs (GA + CA + EA + 5-FU) group. All the data are expressed as mean ± SD (n = 3). (d) CI and DRI values from different combined groups are demonstrated in the heat-maps plotted by R programming, which are assessed by the color bars on the right (→red, values are greater; →blue, values are lower; 5-FU1 and 5-FU2 represent DRIs in the DY + 5-FU and tanins+5-FU groups, respectively; F30–F90, 30% to 90% effective rate).
Mentions: Tannins are the major components in DY (~17% weight of radix)21. Gallic acid (GA) and ellagic acid (EA) are representative hydrolysable tannins, while catechinic acid is one of the oligomeric procyanidins in DY. In this study, a water extract of DY was prepared using an extraction method with good reproducibility (RSD% of GA content = 1.39%, Supplementary Fig. 1a), and the three tannins in the overall extract of DY were separated and identified by HPLC analysis (Fig. 5 and Supplementary Fig. 1b). As determined by HPLC, the three tannins accounted for 3.9% of the total content of the DY extract. To investigate whether GA, CA and EA were the major bioactive constituents of DY that were responsible for the anti-proliferative effects on the CRC cell lines, we evaluated the cytotoxicity of each tannin in CRC cells. No obvious inhibitory activity was observed for any of the tannins (IC50 > 100 μM). Then, we concurrently treated the cells with two or three tannins at same concentrations in 100 μg/ml DY extracts. The cell viabilities were significantly inhibited by the combined tannins (P < 0.05, vs single tannin group), and treatments with two or three tannins showed comparable potency in inhibiting the CRC cells (we chose the combination of three tannins for further research considering its better cooperation). However, the inhibitory effects were lower than that of the overall DY extract (P < 0.05) (Fig. 6a). The dose-effect curves of the three tannins are shown in Fig. 6b. The HCT-116 cells were more sensitive to the tannin treatment than the RKO cells, the growth of which was notably inhibited in a dose-dependent manner. The inhibitory effects of the tannins and the DY extract were comparable at low concentrations (25–50 μg/ml). However, with increased concentrations, the anti-proliferative effects of the tannins turned to be much lower than that of the DY treatment. These results indicate that GA, CA and EA together could effectively inhibit the CRC cells but can’t replace DY.

View Article: PubMed Central - PubMed

ABSTRACT

Sanguisorba officinalis L. radix is a widely used herb called DiYu (DY) in China and has an extensive range of bioactivities, including anti-cancer, anti-inflammatory, and anti-oxidative activities. However, there is little evidence to support its anti-cancer effects against colorectal cancer (CRC). The first-line chemotherapeutic agent 5-fluorouracil (5-FU) is used to treat CRC, but its efficiency is hampered by acquired drug resistance. This study found that a water extract of DY exerted anti-proliferative effects against two CRC cell lines (HCT-116 and RKO), and it sensitized CRC cells to 5-FU therapy by activating a reactive oxygen species (ROS)-mediated, mitochondria-caspase-dependent apoptotic pathway. Co-treatment of DY and 5-FU significantly elevated ROS levels, up-regulated Bax/Bcl-2 ratio and triggered mitochondrial dysfunction, followed by a release of cytochrome c and up-regulation of proteins such as cleaved-caspase-9/3 and cleaved-PARP. Additionally, the induction of autophagy may be involved in mediating synergism of DY in HCT-116 cells. Gallic acid (GA), catechinic acid (CA) and ellagic acid (EA) were identified as the potential chief constituents responsible for the synergistic effects of DY. In conclusion, co-treatment of DY, specifically GA, CA and EA, with 5-FU may be a potential alternative therapeutic strategy for CRC by enhancing an intrinsic apoptotic pathway.

No MeSH data available.