Nutr Res Pract.  2019 Aug;13(4):302-309. 10.4162/nrp.2019.13.4.302.

Carpinus turczaninowii extract modulates arterial inflammatory response: a potential therapeutic use for atherosclerosis

• Son YK
• Yoon SR
• Bang WY
• Bae CH
• Yeo JH
• Yeo R
• An J
• Song J
• Kim OY

• KMID: 2453108
• DOI: http://doi.org/10.4162/nrp.2019.13.4.302

Abstract

BACKGROUND/OBJECTIVES
Vascular inflammation is an important feature in the atherosclerotic process. Recent studies report that leaves and branches of Carpinus turczaninowii (C. turczaninowii) have antioxidant capacity and exert anti-inflammatory effects. However, no study has reported the regulatory effect of C. turczaninowii extract on the arterial inflammatory response. This study therefore investigated modulation of the arterial inflammatory response after exposure to C. turczaninowii extract, using human aortic vascular smooth muscle cells (HAoSMCs).
MATERIALS/METHODS
Scavenging activity of free radicals, total phenolic content (TPC), cell viability, mRNA expressions, and secreted levels of cytokines were measured in LPS-stimulated (10 ng/mL) HAoSMCs treated with the C. turczaninowii extract.
RESULTS
C. turczaninowii extract contains high amounts of TPC (225.6 ± 21.0 mg of gallic acid equivalents/g of the extract), as well as exerts time-and dose-dependent increases in strongly scavenged free radicals (average 14.8 ± 1.97 µg/mL IC50 at 40 min). Cell viabilities after exposure to the extracts (1 and 10 µg/mL) were similar to the viability of non-treated cells. Cytokine mRNA expressions were significantly suppressed by the extracts (1 and 10 µg/mL) at 6 hours (h) after exposure. Interleukin-6 secretion was dose-dependently suppressed 2 h after incubation with the extract, at 1-10 µg/mL in non-stimulated cells, and at 5 and 10 µg/mL in LPS-stimulated cells. Similar patterns were also observed at 24 h after incubation with the extract (at 1-10 µg/mL in non-stimulated cells, and at 10 µg/mL in the LPS-stimulated cells). Soluble intracellular vascular adhesion molecules (sICAM-1) secreted from non-stimulated cells and LPS-stimulated cells were similarly suppressed in a dose-dependent manner after 24 h exposure to the extracts, but not after 2 h. In addition, sICAM-1 concentration after 24 h treatment was positively related to IL-6 levels after 2 h and 24 h exposure (r = 0.418, P = 0.003, and r = 0.524, P < 0.001, respectively).
CONCLUSIONS
This study demonstrates that C. turczaninowii modulates the arterial inflammatory response, and indicates the potential to be applied as a therapeutic use for atherosclerosis.

Keyword

Antioxidants; inflammation; cytokines; atherosclerosis; arteries

MeSH Terms

Antioxidants
Arteries
Atherosclerosis*
Betulaceae*
Cell Survival
Cytokines
Free Radicals
Gallic Acid
Humans
Inflammation
Inhibitory Concentration 50
Interleukin-6
Muscle, Smooth, Vascular
Phenol
RNA, Messenger
Antioxidants
Cytokines
Free Radicals
Gallic Acid
Interleukin-6
Phenol
RNA, Messenger

Figure

• 

  Fig. 1 Time- and dose-dependent DPPH free radical scavenging activities (%) of C. turczaninowii extract. Data are expressed as means ± SD. The experiment includes 4 repeats per condition. C. turczaninowii: Carpinus turczaninowii

• 

  Fig. 2 Cell viability (%) of human aortic vascular smooth muscle cells (HAoSMCs) treated with C. turczaninowii extract. Data are expressed as means ± SD. *P < 0.05, **P < 0.001 compared to CON (non-treated control cells). Cells were treated with various concentrations of C. turczaninowii extract. After 24 h, cell viabilities were measured using the CytoX assay. The experiment includes 4 repeats per condition. C. turczaninowii: Carpinus turczaninowii

• 

  Fig. 3 C. turczaninowii extract modulates mRNA expression of pro-inflammatory cytokines in HAoSMCs. The mRNA levels of IL-6 (A) and TNF-α (B) in HAoSMCs were measured using quantitative real time PCR (qPCR). HAoSMCs were treated with the extract (1 and 10 µg/mL) and incubated for 6 h. β-actin was amplified as an internal control. The values are presented by relative quantity (RQ). Data are expressed as means ± SD, and each experiment includes 4 repeats per conditions. *P < 0.05 compared with CON (non-treated control cells); C. turczaninowii, Carpinus turczaninowii; IL-6, interleukin-6; sICAM-1, soluble intracellular adhesion molecule-1; TNF-α, tumor necrosis factor-α

• 

  Fig. 4 Suppressive effect of C. turczaninowii extract on the productions of IL-6 and sICAM-1 secreted from LPS-stimulated HAoSMCs. HAoSMCs were treated with the extract 15 min after LPS stimulation or not, and then incubated for further 2 h and 24 h. The experiment includes 4 repeats per conditions. Concentrations of IL-6 (A) and sICAM-1 (B) in cell culture supernatants were measured by the ELISA assay. Data are expressed as means ± SD. ¶P < 0.1, *P < 0.05, *P < 0.01, *P < 0.001 compared to non-stimulated CON; †P < 0.05, ‡P < 0.001 compared to LPS-stimulated CON; CON, non-treated control; C. turczaninowii, Carpinus turczaninowii; IL-6, interleukin-6; sICAM-1, soluble intracellular adhesion molecule-1

• 

  Fig. 5 Relationship between IL-6 and sICAM-1 concentrations in the cell culture supernatants from HAoSMCs. HAoSMCs were treated with C. turczaninowii extract (0, 1, 5 and 10 µg/mL) 15 min after LPS stimulation or not, and then incubated for further 2 h and 24 h. IL-6 and sICAM-1 concentrations in the cell culture supernatants were measured by the ELISA assay. Tested by Pearson correlation analysis; r = correlation coefficient. The experiment includes 4 repeats per conditions. C. turczaninowii, Carpinus turczaninowii; IL-6, interleukin-6; sICAM-1, soluble intracellular adhesion molecule-1

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