Korean J Physiol Pharmacol.  2013 Apr;17(2):133-137. 10.4196/kjpp.2013.17.2.133.

Ginsenoside Rg2 Inhibits Lipopolysaccharide-Induced Adhesion Molecule Expression in Human Umbilical Vein Endothelial Cell

• Cho YS
• Kim CH
• Ha TS
• Lee SJ
• Ahn HY

• KMID: 1429384
• DOI: http://doi.org/10.4196/kjpp.2013.17.2.133

Abstract

Vascular cell adhesion molecule 1 (VCAM-1), intercellular adhesion molecule 1 (ICAM-1), P- and E-selectin play a pivotal role for initiation of atherosclerosis. Ginsenoside, a class of steroid glycosides, is abundant in Panax ginseng root, which has been used for prevention of illness in Korea. In this study, we investigated the mechanism(s) by which ginsenoside Rg2 may inhibit VCAM-1 and ICAM-1 expressions stimulated with lipopolysaccharide (LPS) in human umbilical vein endothelial cell (HUVEC). LPS increased VCAM-1 and ICAM-1 expression. Ginsenoside Rg2 prevented LPS-mediated increase of VCAM-1 and ICAM-1 expression. On the other hand, JSH, a nuclear factor kappa B (NF-kappaB) inhibitor, reduced both VCAM-1 and ICAM-1 expression stimulated with LPS. SB202190, inhibitor of p38 mitogen-activated protein kinase (p38 MAPK), and wortmannin, phosphatidylinositol 3-kinase (PI3-kinase) inhibitor, reduced LPS-mediated VCAM-1 but not ICAM-1 expression. PD98059, inhibitor of mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (MEK/ERK) did not affect VCAM-1 and ICAM-1 expression stimulated with LPS. SP600125, inhibitor of c-Jun N-terminal kinase (JNK), reduced LPS-mediated ICAM-1 but not VCAM-1 expression. LPS reduced IkappaBalpha (IkappaBalpha) expression, in a time-dependent manner within 1 hr. Ginsenoside Rg2 prevented the decrease of IkappaBalpha expression stimulated with LPS. Moreover, ginsenoside Rg2 reduced LPS-mediated THP-1 monocyte adhesion to HUVEC, in a concentration-dependent manner. These data provide a novel mechanism where the ginsenoside Rg2 may provide direct vascular benefits with inhibition of leukocyte adhesion into vascular wall thereby providing protection against vascular inflammatory disease.

Keyword

Endothelial cell; Ginsenoside Rg2; ICAM-1; VCAM-1

MeSH Terms

Androstadienes
Anthracenes
Atherosclerosis
E-Selectin
Endothelial Cells
Flavonoids
Ginsenosides
Glycosides
Hand
Humans
I-kappa B Proteins
Imidazoles
Intercellular Adhesion Molecule-1
JNK Mitogen-Activated Protein Kinases
Korea
Leukocytes
Monocytes
NF-kappa B
Panax
Phosphatidylinositol 3-Kinase
Phosphotransferases
Protein Kinases
Pyridines
Umbilical Veins
Vascular Cell Adhesion Molecule-1
Androstadienes
Anthracenes
E-Selectin
Flavonoids
Ginsenosides
Glycosides
I-kappa B Proteins
Imidazoles
Intercellular Adhesion Molecule-1
JNK Mitogen-Activated Protein Kinases
NF-kappa B
Phosphatidylinositol 3-Kinase
Phosphotransferases
Protein Kinases
Pyridines
Vascular Cell Adhesion Molecule-1

Figure

• 

  Fig. 1 Effect of Rg2 on the protein expression levels of adhesion molecules in HUVEC stimulated with LPS. Endothelial cell was treated with 1, 10, 20, 50 and 100µmol/l of Rg2 for 1 hr prior to LPS (1µg/ml) stimulation for 8 hr. Cell extracts were resolved on 8% SDS-polyacrylamide gel and Western blot analysis with the respective primary antibody against VCAM-1 and ICAM-1. β-tubulin was used as an internal control. The bar graph represents the amount of VCAM-1 and ICAM-1 estimated by image scanning and is expressed in arbitrary units. Values are means±SEM of 3 independent experiments. Statistical significance assessed by one-way ANOVA followed by Scheffe post-hoc test for multiple comparisons (**p<0.01, ***p<0.001 vs LPS).

• 

  Fig. 2 Effect of inhibitors of signal transduction pathway on the protein expression levels of adhesion molecules in HUVEC stimulated with LPS. Endothelial cell was treated with 50µmol/l of JSH (NF-κB inhibitor), SB202190 (p38 MAPK inhibitor), PD98059 (MEK/ERK inhibitor), wortmannin (PI3-K inhibitor), and SP600125 (JNK inhibitor) for 1hr prior to LPS (1µg/ml) stimulation for 8 hr. Cell extracts were resolved on 8% SDS-polyacrylamide gel and Western blot analysis with the respective primary antibody against VCAM-1 and ICAM-1. β-tubulin was used as an internal control. The bar graph represents the amount of VCAM-1 and ICAM-1 estimated by image scanning and is expressed in arbitrary units. Values are means±SEM of 3 independent experiments. Statistical significance assessed by one-way ANOVA followed by Scheffe post-hoc test for multiple comparisons (*p<0.05, ***p<0.001 vs LPS).

• 

  Fig. 3 Effect of Rg2 on IκBα expression in HUVEC stimulated with LPS. (A) Time-course of IκBα expression in HUVECs stimulated with LPS (1µg/ml). (B) Effect of Rg2 on time-course of IκBα expression in HUVEC stimulated with LPS. Endothelial cell was treated with 1, 20, and 50µmol/l of Rg2 prior to LPS (1µg/ml) stimulation for 1 hr. Cell extracts were resolved on 12% SDS-polyacrylamide gel and Western blot analysis with the respective primary antibody against IκBα. β-tubulin was used as an internal control. The bar graph represents the amount of IκBα estimated by image scanning and is expressed in arbitrary units. Values are means±SEM of 3 independent experiments. Statistical significance assessed by one-way ANOVA followed by Scheffe post-hoc test for multiple comparisons (A: **p<0.05 vs control; B: *p<0.05, **p<0.01 vs LPS).

• 

  Fig. 4 Effect of Rg2 on THP-1 monocyte adhesion to LPS in HUVEC stimulated with LPS. (A) Endothelial cell was treated with 1, 20, and 50µmol/l of Rg2 prior to LPS (1µg/ml) stimulation for 8 hr. THP-1 cells were labeled with Calcein-AM (5µmol/l) for 30 min. The labeled THP-1 cells were seeded at a density of 5.0×105 cells/well onto endothelial cells treated with the Rg2 and/or LPS and then incubated for 1 hr. Microphotographs (four independent experiments) were obtained using fluorescence microscopy. Magnification ×100. (B) The bar graph represents the cell number of THP-1 monocyte. Values are means±SEM of 3 independent experiments. Statistical significance assessed by one-way ANOVA followed by Scheffe post-hoc test for multiple comparisons (*p<0.05, **p<0.01 vs LPS).

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