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Ethanol extract of Forsythia suspensa root induces apoptosis of esophageal carcinoma cells via the mitochondrial apoptotic pathway.

Zhao L, Yan X, Shi J, Ren F, Liu L, Sun S, Shan B - Mol Med Rep (2014)

Bottom Line: The results revealed that FSREE, rather than Forsythia suspensa ethanolic extracts from the leaf (FSLEE) and fruit (FSFEE) exhibited marked anti-tumor activity towards human esophageal cancer cells.FSREE induced cancer cell apoptosis and growth arrest by downregulating B‑cell lymphoma (Bcl)‑2, Bcl‑extra large and myeloid cell leukemia 1, while upregulating Bcl‑2‑associated X protein, Bcl‑2 antagonist of cell death and phorbol‑12‑myristate‑13‑acetate‑induced protein 1.Furthermore, the anti-cancer activity of FSREE was associated with a decreased level of phosphorylated Janus kinase/signal transducer and activator of transcription 3 and extracellular‑signal‑regulated kinase signaling activity.

View Article: PubMed Central - PubMed

Affiliation: Research Center, The Fourth Hospital of Hebei Medical University, Shijiazhuang, Hebei 050011, P.R. China.

ABSTRACT
Forsythia suspensa root is used in the treatment of fever and jaundice in Traditional Chinese Medicine. In the present study, the anti-tumor activity of the ethanolic extract of Forsythia suspensa root (FSREE) against esophageal carcinoma cells was investigated in vitro and in vivo and its anti-cancer mechanism was examined. The results revealed that FSREE, rather than Forsythia suspensa ethanolic extracts from the leaf (FSLEE) and fruit (FSFEE) exhibited marked anti-tumor activity towards human esophageal cancer cells. FSREE induced cancer cell apoptosis and growth arrest by downregulating B‑cell lymphoma (Bcl)‑2, Bcl‑extra large and myeloid cell leukemia 1, while upregulating Bcl‑2‑associated X protein, Bcl‑2 antagonist of cell death and phorbol‑12‑myristate‑13‑acetate‑induced protein 1. This led to the activation of poly(ADP ribose) polymerase, caspase‑3 and caspase‑9, but not caspase‑8. Furthermore, the anti-cancer activity of FSREE was associated with a decreased level of phosphorylated Janus kinase/signal transducer and activator of transcription 3 and extracellular‑signal‑regulated kinase signaling activity. It was also observed that the levels of cytochrome c were elevated in the cytoplasm, accounting for the loss of mitochondrial membrane potential in the TE‑13 cells upon treatment with FSEER. In addition, FSEER inhibited the growth of esophageal cancer cells in xenograft models and no detectable toxicity was present in the lung or liver tissues. These observations provided further evidence of the anti-tumor effect of FSEER and may be of importance to further examine the potential role of Forsythia suspensa root as a therapeutic agent in esophageal carcinoma therapy.

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(A) Morphological changes of TE-13, TE-1, Eca-109 and Yes-2 cells treated with 0.5 mg/ml FSEER for 48 h. (B) TE-13 cells were treated with 0.251 mg/ml FSEER for 48 h and stained using Giemsa for 10 min. (C) Effects of FSEER on the apoptotic rate of TE-13 cells following a 48 h treatment. Representative histograms of Annexin fluorescein isothiocyanate-stained and propidium iodide-stained cells. *P<0.05, **P<0.01, compared with the control group. (D) Histogram demonstrating increased MMP by 0.5 mg/ml FSEER. *P<0.05, **P<0.01, compared with the control group. FSEER, Forsythia suspensa ethanolic extract of the root; MMP, mitochondrial membrane potential.
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f2-mmr-11-02-0871: (A) Morphological changes of TE-13, TE-1, Eca-109 and Yes-2 cells treated with 0.5 mg/ml FSEER for 48 h. (B) TE-13 cells were treated with 0.251 mg/ml FSEER for 48 h and stained using Giemsa for 10 min. (C) Effects of FSEER on the apoptotic rate of TE-13 cells following a 48 h treatment. Representative histograms of Annexin fluorescein isothiocyanate-stained and propidium iodide-stained cells. *P<0.05, **P<0.01, compared with the control group. (D) Histogram demonstrating increased MMP by 0.5 mg/ml FSEER. *P<0.05, **P<0.01, compared with the control group. FSEER, Forsythia suspensa ethanolic extract of the root; MMP, mitochondrial membrane potential.

Mentions: Giemsa staining and flow cytometry were performed to investigate whether FSEER induced TE-13 cell apoptosis. As shown in Fig. 2A and B, morphological changes observed using microscopy and Giemsa staining revealed that tumor cells exhibited decreased growth, loss of volume, cytoplasm concentration, karyokinesis and deformation to a round appearance following treatment with FSEER (0.5 mg/ml) for 48 h. However, the cells in the control group were observed to maintain a regular appearance, intensive growth and a polygonal shape. Flow cytometry was performed to estimate the rate of apoptosis by quantitative assessment of Annexin V/PI stained TE-13 cells. As shown in Fig. 2C, FSEER treatment increased the number of Annexin V-FITC-positive and PI-negative cells in a dose- and time-dependent manner compared with that in the control group. In order to determine whether FSEER-induced apoptosis of TE-13 cells was mediated through mitochondrial dysfunction, the MMP was measured using the mitochondrial-sensitive dye JC-1. As shown in Fig. 2D, the number of cells exhibiting depolarized mitochondrial membranes was significantly increased in the FSEER (0.25, 0.5, 1.0 mg/ml)-treated cells compared with that in the control group.


Ethanol extract of Forsythia suspensa root induces apoptosis of esophageal carcinoma cells via the mitochondrial apoptotic pathway.

Zhao L, Yan X, Shi J, Ren F, Liu L, Sun S, Shan B - Mol Med Rep (2014)

(A) Morphological changes of TE-13, TE-1, Eca-109 and Yes-2 cells treated with 0.5 mg/ml FSEER for 48 h. (B) TE-13 cells were treated with 0.251 mg/ml FSEER for 48 h and stained using Giemsa for 10 min. (C) Effects of FSEER on the apoptotic rate of TE-13 cells following a 48 h treatment. Representative histograms of Annexin fluorescein isothiocyanate-stained and propidium iodide-stained cells. *P<0.05, **P<0.01, compared with the control group. (D) Histogram demonstrating increased MMP by 0.5 mg/ml FSEER. *P<0.05, **P<0.01, compared with the control group. FSEER, Forsythia suspensa ethanolic extract of the root; MMP, mitochondrial membrane potential.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4262507&req=5

f2-mmr-11-02-0871: (A) Morphological changes of TE-13, TE-1, Eca-109 and Yes-2 cells treated with 0.5 mg/ml FSEER for 48 h. (B) TE-13 cells were treated with 0.251 mg/ml FSEER for 48 h and stained using Giemsa for 10 min. (C) Effects of FSEER on the apoptotic rate of TE-13 cells following a 48 h treatment. Representative histograms of Annexin fluorescein isothiocyanate-stained and propidium iodide-stained cells. *P<0.05, **P<0.01, compared with the control group. (D) Histogram demonstrating increased MMP by 0.5 mg/ml FSEER. *P<0.05, **P<0.01, compared with the control group. FSEER, Forsythia suspensa ethanolic extract of the root; MMP, mitochondrial membrane potential.
Mentions: Giemsa staining and flow cytometry were performed to investigate whether FSEER induced TE-13 cell apoptosis. As shown in Fig. 2A and B, morphological changes observed using microscopy and Giemsa staining revealed that tumor cells exhibited decreased growth, loss of volume, cytoplasm concentration, karyokinesis and deformation to a round appearance following treatment with FSEER (0.5 mg/ml) for 48 h. However, the cells in the control group were observed to maintain a regular appearance, intensive growth and a polygonal shape. Flow cytometry was performed to estimate the rate of apoptosis by quantitative assessment of Annexin V/PI stained TE-13 cells. As shown in Fig. 2C, FSEER treatment increased the number of Annexin V-FITC-positive and PI-negative cells in a dose- and time-dependent manner compared with that in the control group. In order to determine whether FSEER-induced apoptosis of TE-13 cells was mediated through mitochondrial dysfunction, the MMP was measured using the mitochondrial-sensitive dye JC-1. As shown in Fig. 2D, the number of cells exhibiting depolarized mitochondrial membranes was significantly increased in the FSEER (0.25, 0.5, 1.0 mg/ml)-treated cells compared with that in the control group.

Bottom Line: The results revealed that FSREE, rather than Forsythia suspensa ethanolic extracts from the leaf (FSLEE) and fruit (FSFEE) exhibited marked anti-tumor activity towards human esophageal cancer cells.FSREE induced cancer cell apoptosis and growth arrest by downregulating B‑cell lymphoma (Bcl)‑2, Bcl‑extra large and myeloid cell leukemia 1, while upregulating Bcl‑2‑associated X protein, Bcl‑2 antagonist of cell death and phorbol‑12‑myristate‑13‑acetate‑induced protein 1.Furthermore, the anti-cancer activity of FSREE was associated with a decreased level of phosphorylated Janus kinase/signal transducer and activator of transcription 3 and extracellular‑signal‑regulated kinase signaling activity.

View Article: PubMed Central - PubMed

Affiliation: Research Center, The Fourth Hospital of Hebei Medical University, Shijiazhuang, Hebei 050011, P.R. China.

ABSTRACT
Forsythia suspensa root is used in the treatment of fever and jaundice in Traditional Chinese Medicine. In the present study, the anti-tumor activity of the ethanolic extract of Forsythia suspensa root (FSREE) against esophageal carcinoma cells was investigated in vitro and in vivo and its anti-cancer mechanism was examined. The results revealed that FSREE, rather than Forsythia suspensa ethanolic extracts from the leaf (FSLEE) and fruit (FSFEE) exhibited marked anti-tumor activity towards human esophageal cancer cells. FSREE induced cancer cell apoptosis and growth arrest by downregulating B‑cell lymphoma (Bcl)‑2, Bcl‑extra large and myeloid cell leukemia 1, while upregulating Bcl‑2‑associated X protein, Bcl‑2 antagonist of cell death and phorbol‑12‑myristate‑13‑acetate‑induced protein 1. This led to the activation of poly(ADP ribose) polymerase, caspase‑3 and caspase‑9, but not caspase‑8. Furthermore, the anti-cancer activity of FSREE was associated with a decreased level of phosphorylated Janus kinase/signal transducer and activator of transcription 3 and extracellular‑signal‑regulated kinase signaling activity. It was also observed that the levels of cytochrome c were elevated in the cytoplasm, accounting for the loss of mitochondrial membrane potential in the TE‑13 cells upon treatment with FSEER. In addition, FSEER inhibited the growth of esophageal cancer cells in xenograft models and no detectable toxicity was present in the lung or liver tissues. These observations provided further evidence of the anti-tumor effect of FSEER and may be of importance to further examine the potential role of Forsythia suspensa root as a therapeutic agent in esophageal carcinoma therapy.

Show MeSH
Related in: MedlinePlus