Nutr Res Pract.  2016 Jun;10(3):259-264. 10.4162/nrp.2016.10.3.259.

Compound K attenuates stromal cell-derived growth factor 1 (SDF-1)-induced migration of C6 glioma cells

• Kim H
• Roh HS
• Kim JE
• Park SD
• Park WH
• Moon JY

• KMID: 2342127
• DOI: http://doi.org/10.4162/nrp.2016.10.3.259

Abstract

BACKGROUND/OBJECTIVES
Stromal cell-derived growth factor 1 (SDF-1), also known as chemokine ligand 12, and chemokine receptor type 4 are involved in cancer cell migration. Compound K (CK), a metabolite of protopanaxadiol-type ginsenoside by gut microbiota, is reported to have therapeutic potential in cancer therapy. However, the inhibitory effect of CK on SDF-1 pathway-induced migration of glioma has not yet been established.
MATERIALS/METHODS
Cytotoxicity of CK in C6 glioma cells was determined using an EZ-Cytox cell viability assay kit. Cell migration was tested using the wound healing and Boyden chamber assay. Phosphorylation levels of protein kinase C (PKC)α and extracellular signal-regulated kinase (ERK) were measured by western blot assay, and matrix metallopeptidases (MMP) were measured by gelatin-zymography analysis.
RESULTS
CK significantly reduced the phosphorylation of PKCα and ERK1/2, expression of MMP9 and MMP2, and inhibited the migration of C6 glioma cells under SDF-1-stimulated conditions.
CONCLUSIONS
CK is a cell migration inhibitor that inhibits C6 glioma cell migration by regulating its downstream signaling molecules including PKCα, ERK1/2, and MMPs.

Keyword

Panax ginseng; compound K; SDF-1; anti-tumor; C6 glioma

MeSH Terms

Blotting, Western
Cell Movement
Cell Survival
Gastrointestinal Microbiome
Glioma*
Matrix Metalloproteinases
Metalloproteases
Panax
Phosphorylation
Phosphotransferases
Protein Kinase C
Wound Healing
Matrix Metalloproteinases
Metalloproteases
Phosphotransferases
Protein Kinase C

Figure

• 

  Fig. 1 Effect of CK on viability of C6 glioma cells. (A) Chemical structure of compound K (CK), [20-,O-β-(D-glucopyranosyl)-20(S)-protopanaxadiol]. (B) C6 glioma cells were incubated in the presence or absence of CK (0.03 µM to 10 µM) in serum-depleted medium for 24 h. Bar graph indicates the effect of CK on C6 glioma cell viability. The values were expressed as mean ± SE of three independent experiments. *P < 0.05 vs. untreated C6 glioma cells.

• 

  Fig. 2 Effect of CK on SDF-1-induced migration of C6 glioma cells in the wound-healing assay. (A) Microphotographs of SDF-1-stimulated C6 glioma cells, the white dotted lines in each result indicate the first scratch (upper panel) at 0 h and the altered scratch at 24 h (lower panel). No-treatment or treatment with CK (0.1 µM -1 µM) affected the rate of migration as indicated by scratch-wound assay. (B) The graph indicates the cell migration ratio of CK-treated and SDF-1-stimulated C6 glioma cells compared with the control group. The values are expressed as mean ± SE of three independent experiments. *P < 0.05 vs. SDF-1 alone.

• 

  Fig. 3 Effect of CK on SDF-1-induced migration of C6 glioma cells. (A) Microphotographs of SDF-1-induced migration of C6 glioma cells as determined by Boyden chamber assay. C6 glioma cells were treated with vehicle or CK (0.1 µM -1 µM) for 90 min in SDF-1 containing medium. (B) Quantification of migration ratio of SDF-1-stimulated C6 glioma cells. The graph indicates CK-mediated inhibition of SDF-1-stimulated C6 glioma cell migration compared to the untreated group. The values are expressed as mean ± SE of three independent experiments. *P < 0.05 vs. SDF-1 alone.

• 

  Fig. 4 Effect of CK on phosphorylation of PKCα and ERK1/2 in SDF-1-stimulated C6 glioma cells. C6 glioma cells were pretreated with CK (0.1 µM -1 µM), followed by stimulation with SDF-1 (100 ng/ml) for an additional 24 h. (A) Expression levels of phospho-PKCα (P-PKCα), PKCα (T-PKCα), phospho-ERK1/2 (P-ERK1/2), and ERK1/2 (T-ERK1/2) were analyzed by western blotting. (B-C) The intensity of phosphorylation of PKCα and ERK1/2 was quantified by densitometry. The values were expressed as mean ± SE of three independent experiments. *P < 0.05 vs. SDF-1 alone.

• 

  Fig. 5 The effect of CK on MMPs in SDF-1 stimulated C6 glioma cells. C6 glioma cells were incubated in serum-depleted medium for 24 h. Cells were pre-treated with CK (0.03 µM to 1 µM) for 1 h, followed by treatment with SDF-1 for 23 h. These supernatants were analyzed by gelatin-zymography analysis. (A) Expression of MMP9 and MMP2 in zymorgraphy gel. (B-C) The graph indicates the quantification of MMP2 and MMP9 by densitometry. The values are expressed as mean ± SE of three independent experiments. *P < 0.05 vs. SDF-1 alone.

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