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Journal of Chinese Integrative Medicine ›› 2012, Vol. 10 ›› Issue (10): 1155-.doi: 10.3736/jcim20121013

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Danshensu inhibits acetaldehyde-induced proliferation and activation of hepatic stellate cell-T6

Li Zhang1, Tao Wu2, Jun-ming Chen3, Li-li Yang1, Hai-yan Song1, Guang Ji1()   

  1. 1. Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
    2. Center of Chinese Medicine Therapy and Systems Biology, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
    3. Department of Traditional Chinese Medicine, Shanghai Sixth People’s Hospital, Shanghai Jiaotong University, Shanghai 200233, China;
  • Received:2012-04-09 Accepted:2012-05-22 Online:2012-10-20 Published:2018-10-15
  • Contact: Guang Ji E-mail:jiliver@vip.sina.com

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Objective: To evaluate the effects of danshensu, the main component of the extract of Chinese medicineSalvia miltiorrhiza, on the proliferation and activation of hepatic stellate cells (HSCs). .

Methods: The activation of HSC-T6 was induced by exposure to acetaldehyde. In the meantime, different doses of danshensu were added to the culture medium. After 24 h of treatment with danshensu in acetaldehyde, the viability of HSC-T6 cells was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, the cell cycle was determined through flow cytometry, and the gene transcription levels of plasminogen activator inhibitor-1 (PAI-1), transforming growth factor-β1 (TGF-β1), urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-2 (MMP-2) were analyzed by real-time quantitative polymerase chain reaction.RESULTS: The proliferation of HSCs induced by 200 μmol/L acetaldehyde could be significantly inhibited by danshensu, and the percentage of HSCs in S phase was significantly increased as compared with the control cells (P<0.05), which were respectively evidenced by MTT assay and flow cytometry. Danshensu down-regulated the mRNA expression of TGF-β1 and PAI-1 and up-regulated the uPA transcription level (P<0.01), while the transcription level of MMP-2 was not significantly affected in HSC-T6.

Conclusion: Danshensu can inhibit the proliferation and activation of HSC-T6, as well as regulate some cytokines involved in extracellular matrix accumulation, which offers a potential therapeutic alternative for liver fibrosis.

Key words: danshensu, hepatocytes, cell proliferation, extracellular matrix, in vitro

"

Primer Primer sequence
PAI-1 Forward 5′-GTGGTTCGGCACAATCCAACAGAG-3′
Reverse 5′-GCAATGGAGGACGATACAAGAGGT-3′
uPA Forward 5′-AAGGTCTGGGATACTAGAGCC-3′
Reverse 5′-CAGGATATCCTCAGTCTCCACC-3′
TGF-β1 Forward 5′-GCCCTGGACACCAACTATTGC-3′
Reverse 5′-GCTGCACTTGCAGGAGCGCAC-3′
MMP-2 Forward 5′-ACTCCCACTTTGATGACGAC-3′
Reverse 5′-GAAGAGGAAAGGGAACTTGC-3′
GAPDH Forward 5′-GGTCGGAGTCAACGGATTTG-3′
Reverse 5′-ATGAGCCCCCAGCCTTCTCCA-3′

Figure 1

Viability of HSC-T6 cells affected by different doses of acetaldehyde and danshensuA: line graph showing the percentage of viable cells subjected to acetaldehyde (0, 100, 200, and 300 μmol/L); B: line graph showing the percentage of viable cells exposed to danshensu (0, 150, 250, 615 and 1 000 μmol/L) respectively. Values were measured in three independent experiments done in triplicate and mean ± standard error was plotted. HSC: hepatic stellate cell. "

Figure 2

Effects of Danshensu on the cell cycle of HSC-T6 HSC-T6 cultured with DMEM as control, and cells cultured with 200 μmol/L acetaldehyde as model. Cells were exposed to 200 μmol/L acetaldehyde with danshensu in different concentrations (100, 125 and 150 μmol/L) for 24 h. Cells were suspended, fixed and stained with PI, then were analyzed by flow cytometry. HSC: hepatic stellate cell; DMEM: Dulbecco’s Modified Eagle’s Medium; D: danshensu; PI: propidium iodide."

Figure 3

Effects of danshensu on PAI-1, TGF-β1, uPA and MMP-2 expressionGene transcription levels were measured by real-time quantitative polymerase chain reaction, with GAPDH expression level as the native control. Experiment for each sample was carried out in triplicate and data were expressed as mean±standard error. a: PAI-1 mRNA expression; b: TGF-β1 mRNA expression; c: cytokine uPA gene expression; d: cytokine MMP-2 gene expression. **P<0.01, vs control group; △△P<0.01, vs model group. Ace: acetaldehyde; D: danshensu; PAI-1: plasminogen activator inhibitor-1; uPA: urokinase-type plasminogen activator; MMP-2: matrix metalloproteinase-2; TGF-β1: transforming growth factor-β1; GAPDH: glyceraldehyde-3-phosphate dehydrogenase."

Figure 4

The chemical structure of danshensu"

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