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Morin, a flavonoid from moraceae, induces apoptosis by induction of BAD protein in human leukemic cells.

Park C, Lee WS, Go SI, Nagappan A, Han MH, Hong SH, Kim GS, Kim GY, Kwon TK, Ryu CH, Shin SC, Choi YH - Int J Mol Sci (2014)

Bottom Line: The apoptotic activity of morin was significantly attenuated by Bcl-2 augmentation.In conclusion, morin induced caspase-dependent apoptosis through an intrinsic pathway by upregulating BAD proteins.In addition, Bcl-2 protein expression is also important in morin-induced apoptosis of U937 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, College of Natural Sciences, Dongeui University, Busan 614-714, Korea. parkch@deu.ac.kr.

ABSTRACT
Evidence suggests that phytochemicals can safely modulate cancer cell biology and induce apoptosis. Here, we investigated the anti-cancer activity of morin, a flavone originally isolated from members of the Moraceae family in human leukemic cells, focusing on apoptosis. An anti-cancer effect of morin was screened with several human leukemic cell lines. U937 cells were most sensitive to morin, where it induced caspase-dependent apoptosis in a dose-dependent manner. It also induced loss of MMP (ΔΨm) along with cytochrome c release, down-regulated Bcl-2 protein, and up-regulated BAX proteins. The apoptotic activity of morin was significantly attenuated by Bcl-2 augmentation. In conclusion, morin induced caspase-dependent apoptosis through an intrinsic pathway by upregulating BAD proteins. In addition, Bcl-2 protein expression is also important in morin-induced apoptosis of U937 cells. This study provides evidence that morin might have anticancer properties in human leukemic cells.

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Activation of caspases and subsequent cleavage of PARP during the morin-induced apoptosis in U937 cells. U937 cells were incubated at indicated concentrations of morin for 48 h. (A) Western blot analysis for the effects of morin on the caspase activation and PARP cleavage. The membranes were probed with the anti-caspase-3, anti-caspase-8, anti-caspase-9 and anti-PARP antibodies. The expression of the indicated proteins were measured by densitometry and expressed as average relative ratio compared to actin, from two or three different experiments and (B) In vitro assay for caspase-3, -8 and -9 activity, which uses DEVD-pNA, IETD-pNA and LEHD-pNA as substrates, respectively. The released fluorescent products were measured. Data are expressed as mean ± SD of three independent experiments. (*p < 0.05 vs. control).
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ijms-16-00645-f002: Activation of caspases and subsequent cleavage of PARP during the morin-induced apoptosis in U937 cells. U937 cells were incubated at indicated concentrations of morin for 48 h. (A) Western blot analysis for the effects of morin on the caspase activation and PARP cleavage. The membranes were probed with the anti-caspase-3, anti-caspase-8, anti-caspase-9 and anti-PARP antibodies. The expression of the indicated proteins were measured by densitometry and expressed as average relative ratio compared to actin, from two or three different experiments and (B) In vitro assay for caspase-3, -8 and -9 activity, which uses DEVD-pNA, IETD-pNA and LEHD-pNA as substrates, respectively. The released fluorescent products were measured. Data are expressed as mean ± SD of three independent experiments. (*p < 0.05 vs. control).

Mentions: Next, we determined whether morin-induced apoptosis was caspase-dependent. Western blotting analyses revealed that morin activated procaspase-3, procaspase-8, and procaspase-9 in a dose-dependent manner (Figure 2A). Morin also induced the cleavage of PARP, β-catenin, and PLCγ1 which are the substrates of caspases (Figure 2A). We next performed capase activity assays. Morin activated caspase-3 and caspase-9 rather than capase-8 in a dose-dependent manner (Figure 2B). This finding indicates that morin induced capase-3 and caspase-9, which are associated with mitochondria-mediated apoptosis. We confirmed the finding with a caspase-3 inhibitor, z-DEVD-fluoromethylketone (fmk). MTT and DNA fragementation assay revealed that z-DEVD-fmk significantly reduced morin-induced cell death (Figure 3A,B). In addition, Annexin V staining also revealed that z-DEVD-fmk significantly reduced morin-induced cell death Figure 3C). These findings thus suggest that morin induces caspase-dependent apoptosis.


Morin, a flavonoid from moraceae, induces apoptosis by induction of BAD protein in human leukemic cells.

Park C, Lee WS, Go SI, Nagappan A, Han MH, Hong SH, Kim GS, Kim GY, Kwon TK, Ryu CH, Shin SC, Choi YH - Int J Mol Sci (2014)

Activation of caspases and subsequent cleavage of PARP during the morin-induced apoptosis in U937 cells. U937 cells were incubated at indicated concentrations of morin for 48 h. (A) Western blot analysis for the effects of morin on the caspase activation and PARP cleavage. The membranes were probed with the anti-caspase-3, anti-caspase-8, anti-caspase-9 and anti-PARP antibodies. The expression of the indicated proteins were measured by densitometry and expressed as average relative ratio compared to actin, from two or three different experiments and (B) In vitro assay for caspase-3, -8 and -9 activity, which uses DEVD-pNA, IETD-pNA and LEHD-pNA as substrates, respectively. The released fluorescent products were measured. Data are expressed as mean ± SD of three independent experiments. (*p < 0.05 vs. control).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4307266&req=5

ijms-16-00645-f002: Activation of caspases and subsequent cleavage of PARP during the morin-induced apoptosis in U937 cells. U937 cells were incubated at indicated concentrations of morin for 48 h. (A) Western blot analysis for the effects of morin on the caspase activation and PARP cleavage. The membranes were probed with the anti-caspase-3, anti-caspase-8, anti-caspase-9 and anti-PARP antibodies. The expression of the indicated proteins were measured by densitometry and expressed as average relative ratio compared to actin, from two or three different experiments and (B) In vitro assay for caspase-3, -8 and -9 activity, which uses DEVD-pNA, IETD-pNA and LEHD-pNA as substrates, respectively. The released fluorescent products were measured. Data are expressed as mean ± SD of three independent experiments. (*p < 0.05 vs. control).
Mentions: Next, we determined whether morin-induced apoptosis was caspase-dependent. Western blotting analyses revealed that morin activated procaspase-3, procaspase-8, and procaspase-9 in a dose-dependent manner (Figure 2A). Morin also induced the cleavage of PARP, β-catenin, and PLCγ1 which are the substrates of caspases (Figure 2A). We next performed capase activity assays. Morin activated caspase-3 and caspase-9 rather than capase-8 in a dose-dependent manner (Figure 2B). This finding indicates that morin induced capase-3 and caspase-9, which are associated with mitochondria-mediated apoptosis. We confirmed the finding with a caspase-3 inhibitor, z-DEVD-fluoromethylketone (fmk). MTT and DNA fragementation assay revealed that z-DEVD-fmk significantly reduced morin-induced cell death (Figure 3A,B). In addition, Annexin V staining also revealed that z-DEVD-fmk significantly reduced morin-induced cell death Figure 3C). These findings thus suggest that morin induces caspase-dependent apoptosis.

Bottom Line: The apoptotic activity of morin was significantly attenuated by Bcl-2 augmentation.In conclusion, morin induced caspase-dependent apoptosis through an intrinsic pathway by upregulating BAD proteins.In addition, Bcl-2 protein expression is also important in morin-induced apoptosis of U937 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, College of Natural Sciences, Dongeui University, Busan 614-714, Korea. parkch@deu.ac.kr.

ABSTRACT
Evidence suggests that phytochemicals can safely modulate cancer cell biology and induce apoptosis. Here, we investigated the anti-cancer activity of morin, a flavone originally isolated from members of the Moraceae family in human leukemic cells, focusing on apoptosis. An anti-cancer effect of morin was screened with several human leukemic cell lines. U937 cells were most sensitive to morin, where it induced caspase-dependent apoptosis in a dose-dependent manner. It also induced loss of MMP (ΔΨm) along with cytochrome c release, down-regulated Bcl-2 protein, and up-regulated BAX proteins. The apoptotic activity of morin was significantly attenuated by Bcl-2 augmentation. In conclusion, morin induced caspase-dependent apoptosis through an intrinsic pathway by upregulating BAD proteins. In addition, Bcl-2 protein expression is also important in morin-induced apoptosis of U937 cells. This study provides evidence that morin might have anticancer properties in human leukemic cells.

Show MeSH
Related in: MedlinePlus