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Potent effects of dioscin against liver fibrosis.

Zhang X, Han X, Yin L, Xu L, Qi Y, Xu Y, Sun H, Lin Y, Liu K, Peng J - Sci Rep (2015)

Bottom Line: Dioscin effectively inhibited the cell viabilities of HSC-T6, LX-2 and primary rat hepatic stellate cells (HSCs), but not hepatocytes.Furthermore, dioscin markedly increased peroxisome proliferator activated receptor-γ (PPAR-γ) expression and significantly reduced a-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), collagen α1 (I) (COL1A1) and collagen α1 (III) (COL3A1) levels in vitro.Dioscin facilitated matrix degradation, and exhibited hepatoprotective effects through the attenuation of oxidative stress and inflammation, in addition to exerting anti-fibrotic effects through the modulation of the TGF-β1/Smad, Wnt/β-catenin, mitogen-activated protein kinase (MAPK) and mitochondrial signaling pathways, which triggered the senescence of activated HSCs.

View Article: PubMed Central - PubMed

Affiliation: College of Pharmacy, Dalian Medical University, Western 9 Lvshunnan Road, Dalian 116044, China.

ABSTRACT
We previously reported the promising effects of dioscin against liver injury, but its effect on liver fibrosis remains unknown. The present work investigated the activities of dioscin against liver fibrosis and the underlying molecular mechanisms. Dioscin effectively inhibited the cell viabilities of HSC-T6, LX-2 and primary rat hepatic stellate cells (HSCs), but not hepatocytes. Furthermore, dioscin markedly increased peroxisome proliferator activated receptor-γ (PPAR-γ) expression and significantly reduced a-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), collagen α1 (I) (COL1A1) and collagen α1 (III) (COL3A1) levels in vitro. Notably, dioscin inhibited HSCs activation and induced apoptosis in activated HSCs. In vivo, dioscin significantly improved body weight and hydroxylproline, laminin, α-SMA, TGF-β1, COL1A1 and COL3A1 levels, which were confirmed by histopathological assays. Dioscin facilitated matrix degradation, and exhibited hepatoprotective effects through the attenuation of oxidative stress and inflammation, in addition to exerting anti-fibrotic effects through the modulation of the TGF-β1/Smad, Wnt/β-catenin, mitogen-activated protein kinase (MAPK) and mitochondrial signaling pathways, which triggered the senescence of activated HSCs. In conclusion, dioscin exhibited potent effects against liver fibrosis through the modulation of multiple targets and signaling pathways and should be developed as a novel candidate for the treatment of liver fibrosis in the future.

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Dioscin induced apoptosis of HSC-T6, LX2 and primary HSCs.(A) Effects of dioscin on apoptosis of HSC-T6, LX2 and primary HSCs based on AO/EB staining, which were treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h (200×, final magnification). (B) After being treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, HSC-T6, LX2 and primary HSCs were stained with annexin V/PI, and then analyzed by flow cytometry for quantitative detection of cell apoptosis. (C) HSC-T6, LX2 and primary HSCs were incubated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, then the cells were dual-stained for TUNEL (green) and α-SMA (red) (original magnification 200×).
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f2: Dioscin induced apoptosis of HSC-T6, LX2 and primary HSCs.(A) Effects of dioscin on apoptosis of HSC-T6, LX2 and primary HSCs based on AO/EB staining, which were treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h (200×, final magnification). (B) After being treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, HSC-T6, LX2 and primary HSCs were stained with annexin V/PI, and then analyzed by flow cytometry for quantitative detection of cell apoptosis. (C) HSC-T6, LX2 and primary HSCs were incubated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, then the cells were dual-stained for TUNEL (green) and α-SMA (red) (original magnification 200×).

Mentions: Dioscin caused the activated HSCs to undergo apoptosis based on AO/EB fluorescent staining (Figure 2A) and flow cytometry assay (Figure 2B). In the dual-staining for α-SMA and TUNEL, α-SMA-positive cells were obviously decreased and the numbers of TUNEL-positive cells were significantly increased by dioscin (Figure 2C). In contrast to the activated HSCs, dioscin hardly induced the apoptosis in the quiescent HSCs following incubation with 5.0 14μg/mL of diosicn for 24 14h. These results clearly indicated that dioscin induced apoptosis in activated, but not in quiescent HSCs in vitro (Supplemental Figure 2F). Dioscin- treated HSC-T6, LX2 and primary HSCs exhibited cell shrinkage and a loss of the originally confluent monolayer (Supplemental Figure 3). In addition, single cell gel electrophoresis evaluated the extent of dioscin-induced cellular DNA damage. Supplemental Figure 4A demonstrates that nuclei were intact and round and that the DNA remained within the nuclear matrix in control cells. However, obvious DNA damage was observed in dioscin-treated cells. Furthermore, HSCs cells treated with dioscin (5.0 14μg/ml) exhibited fewer cell surface microvilli, chromatin condensation, shrinkage of the cell nuclei and dilated endoplasmic reticulums in apoptotic cells on TEM assays (Supplemental Figure 4B).


Potent effects of dioscin against liver fibrosis.

Zhang X, Han X, Yin L, Xu L, Qi Y, Xu Y, Sun H, Lin Y, Liu K, Peng J - Sci Rep (2015)

Dioscin induced apoptosis of HSC-T6, LX2 and primary HSCs.(A) Effects of dioscin on apoptosis of HSC-T6, LX2 and primary HSCs based on AO/EB staining, which were treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h (200×, final magnification). (B) After being treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, HSC-T6, LX2 and primary HSCs were stained with annexin V/PI, and then analyzed by flow cytometry for quantitative detection of cell apoptosis. (C) HSC-T6, LX2 and primary HSCs were incubated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, then the cells were dual-stained for TUNEL (green) and α-SMA (red) (original magnification 200×).
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Related In: Results  -  Collection

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f2: Dioscin induced apoptosis of HSC-T6, LX2 and primary HSCs.(A) Effects of dioscin on apoptosis of HSC-T6, LX2 and primary HSCs based on AO/EB staining, which were treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h (200×, final magnification). (B) After being treated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, HSC-T6, LX2 and primary HSCs were stained with annexin V/PI, and then analyzed by flow cytometry for quantitative detection of cell apoptosis. (C) HSC-T6, LX2 and primary HSCs were incubated with 5.0 14μg/ml of dioscin for 12, 24 and 48 14h, or treated with different concentrations of dioscin (1.25, 2.5 and 5.0 14μg/ml) for 24 14h, then the cells were dual-stained for TUNEL (green) and α-SMA (red) (original magnification 200×).
Mentions: Dioscin caused the activated HSCs to undergo apoptosis based on AO/EB fluorescent staining (Figure 2A) and flow cytometry assay (Figure 2B). In the dual-staining for α-SMA and TUNEL, α-SMA-positive cells were obviously decreased and the numbers of TUNEL-positive cells were significantly increased by dioscin (Figure 2C). In contrast to the activated HSCs, dioscin hardly induced the apoptosis in the quiescent HSCs following incubation with 5.0 14μg/mL of diosicn for 24 14h. These results clearly indicated that dioscin induced apoptosis in activated, but not in quiescent HSCs in vitro (Supplemental Figure 2F). Dioscin- treated HSC-T6, LX2 and primary HSCs exhibited cell shrinkage and a loss of the originally confluent monolayer (Supplemental Figure 3). In addition, single cell gel electrophoresis evaluated the extent of dioscin-induced cellular DNA damage. Supplemental Figure 4A demonstrates that nuclei were intact and round and that the DNA remained within the nuclear matrix in control cells. However, obvious DNA damage was observed in dioscin-treated cells. Furthermore, HSCs cells treated with dioscin (5.0 14μg/ml) exhibited fewer cell surface microvilli, chromatin condensation, shrinkage of the cell nuclei and dilated endoplasmic reticulums in apoptotic cells on TEM assays (Supplemental Figure 4B).

Bottom Line: Dioscin effectively inhibited the cell viabilities of HSC-T6, LX-2 and primary rat hepatic stellate cells (HSCs), but not hepatocytes.Furthermore, dioscin markedly increased peroxisome proliferator activated receptor-γ (PPAR-γ) expression and significantly reduced a-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), collagen α1 (I) (COL1A1) and collagen α1 (III) (COL3A1) levels in vitro.Dioscin facilitated matrix degradation, and exhibited hepatoprotective effects through the attenuation of oxidative stress and inflammation, in addition to exerting anti-fibrotic effects through the modulation of the TGF-β1/Smad, Wnt/β-catenin, mitogen-activated protein kinase (MAPK) and mitochondrial signaling pathways, which triggered the senescence of activated HSCs.

View Article: PubMed Central - PubMed

Affiliation: College of Pharmacy, Dalian Medical University, Western 9 Lvshunnan Road, Dalian 116044, China.

ABSTRACT
We previously reported the promising effects of dioscin against liver injury, but its effect on liver fibrosis remains unknown. The present work investigated the activities of dioscin against liver fibrosis and the underlying molecular mechanisms. Dioscin effectively inhibited the cell viabilities of HSC-T6, LX-2 and primary rat hepatic stellate cells (HSCs), but not hepatocytes. Furthermore, dioscin markedly increased peroxisome proliferator activated receptor-γ (PPAR-γ) expression and significantly reduced a-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), collagen α1 (I) (COL1A1) and collagen α1 (III) (COL3A1) levels in vitro. Notably, dioscin inhibited HSCs activation and induced apoptosis in activated HSCs. In vivo, dioscin significantly improved body weight and hydroxylproline, laminin, α-SMA, TGF-β1, COL1A1 and COL3A1 levels, which were confirmed by histopathological assays. Dioscin facilitated matrix degradation, and exhibited hepatoprotective effects through the attenuation of oxidative stress and inflammation, in addition to exerting anti-fibrotic effects through the modulation of the TGF-β1/Smad, Wnt/β-catenin, mitogen-activated protein kinase (MAPK) and mitochondrial signaling pathways, which triggered the senescence of activated HSCs. In conclusion, dioscin exhibited potent effects against liver fibrosis through the modulation of multiple targets and signaling pathways and should be developed as a novel candidate for the treatment of liver fibrosis in the future.

Show MeSH
Related in: MedlinePlus