Ann Dermatol.  2012 Feb;24(1):16-21. 10.5021/ad.2012.24.1.16.

Cedrol Enhances Extracellular Matrix Production in Dermal Fibroblasts in a MAPK-Dependent Manner

• Jin MH
• Park SG
• Hwang YL
• Lee MH
• Jeong NJ
• Roh SS
• Lee Y
• Kim CD
• Lee JH

• KMID: 2156850
• DOI: http://doi.org/10.5021/ad.2012.24.1.16

Abstract

BACKGROUND
The extracellular matrix (ECM) produced by dermal fibroblasts supports skin structure, and degradation and/or reduced production of ECM are the main causes of wrinkle formation.
OBJECTIVE
The aim of this study was to identify the active ingredient that enhances ECM production in dermal fibroblasts.
METHODS
Polarity-based fractionation was used to isolate the active ingredient from natural extracts, and the effects of cedrol (isolated from Pterocarpus indicusirginia) on ECM production in cultured human dermal fibroblasts was investigated by reverse transcription-polymerase chain reaction, enzyme linked immunosorbent assay, and Western blot analysis.
RESULTS
Cedrol accelerated fibroblast growth in a dose-dependent manner and increased the production of type 1 collagen and elastin. Phosphorylation of p42/44 extracellular signal-regulated kinase, p38 mitogen-activated protein kinase, and Akt was markedly increased by cedrol, indicating that enhanced ECM production is linked to activation of intracellular signaling cascades.
CONCLUSION
These results indicate that cedrol stimulates ECM production, with possible applications to the maintenance of skin texture.

Keyword

Cedrol; Collagen; Fibroblasts

MeSH Terms

Blotting, Western
Collagen
Collagen Type I
Elastin
Enzyme-Linked Immunosorbent Assay
Extracellular Matrix
Fibroblasts
Humans
Phosphorylation
Phosphotransferases
Protein Kinases
Pterocarpus
Skin
Terpenes
Collagen
Collagen Type I
Elastin
Phosphotransferases
Protein Kinases
Terpenes

Figure

• 

  Fig. 1 (A) Isolation of cedrol from Pterocarpus indicus. (B) Chemical structure of cedrol. Cedrol was isolated from the lignum of P. indicusirginia using bioassay-directed fractionation. After solvent fractionation, two applications of column chromatography followed by recrystallization were used to isolate the active compound. The purity of cedrol was confirmed to be ~99%, as determined by high performance liquid chromatography, nuclear magnetic resonance, and electrospray ionization mass spectrometry studies.

• 

  Fig. 2 Effect of cedrol on growth of dermal fibroblasts. Cells were treated with cedrol at the indicated concentrations for 2 d in the presence of [3H]thymidine. Radioactivity was measured using a liquid scintillation counter. Results are shown as a percentage of a control±the standard deviation (*p<0.05 vs. control).

• 

  Fig. 3 Effect of cedrol on collagen and elastin production in dermal fibroblasts. (A) Cells were treated with cedrol at the indicated concentrations for 2 d. A conditioned medium was collected, and the amount of secreted procollagen type 1 was measured using an enzyme linked immunosorbent assay. Results are shown as a percentage of a control±the standard deviation (*p<0.05 vs. control). (B) Cellular proteins were harvested and the protein levels of collagen type 1 α1 and elastin were verified using Western blot analysis. (C) The expression levels of the COL1A1 and COL1A2 genes were determined using reverse transcription-polymerase chain reaction. CTL: control.

• 

  Fig. 4 Effect of cedrol on intracellular signaling pathways. Cells were treated with cedrol for the indicated times. Cellular proteins were prepared and phosphorylation of ERK, p38 MAPK, Akt, and Smad 2 and 3 was determined using Western blot analysis. CTL: control, ERK: extracellular signal-regulated kinase, MAPK: mitogen-activated protein kinase.

Reference

1. Pieraggi MT, Bouissou H, Angelier C, Uhart D, Magnol JP, Kokolo J. The fibroblast. Ann Pathol. 1985. 5:65–76.

2. Uitto J. Connective tissue biochemistry of the aging dermis. Age-related alterations in collagen and elastin. Dermatol Clin. 1986. 4:433–446.

3. Yada Y, Sadachi H, Nagashima Y, Suzuki T. Overseas survey of the effect of cedrol on the autonomic nervous system in three countries. J Physiol Anthropol. 2007. 26:349–354.

4. Umeno K, Hori E, Tsubota M, Shojaku H, Miwa T, Nagashima Y, et al. Effects of direct cedrol inhalation into the lower airway on autonomic nervous activity in totally laryngectomized subjects. Br J Clin Pharmacol. 2008. 65:188–196.

5. Joseph-Nathan P, Santillan RL, Gutiérrez A. Gutierrez, Carbon-13 NMR. Study of Cedrol, 6-Isocedrol, and α-Cedrene. J Nat Prod. 1984. 47:924–933.

6. Roh SS, Lee MH, Hwang YL, Song HH, Jin MH, Park SG, et al. Stimulation of the extracellular matrix production in dermal fibroblasts by velvet antler extract. Ann Dermatol. 2010. 22:173–179.

7. Ignotz RA, Massagué J. Transforming growth factor-beta stimulates the expression of fibronectin and collagen and their incorporation into the extracellular matrix. J Biol Chem. 1986. 261:4337–4345.

8. Lim IJ, Phan TT, Tan EK, Nguyen TT, Tran E, Longaker MT, et al. Synchronous activation of ERK and phosphatidylinositol 3-kinase pathways is required for collagen and extracellular matrix production in keloids. J Biol Chem. 2003. 278:40851–40858.

9. Lee DJ, Rosenfeldt H, Grinnell F. Activation of ERK and p38 MAP kinases in human fibroblasts during collagen matrix contraction. Exp Cell Res. 2000. 257:190–197.

10. Jinnin M. Mechanisms of skin fibrosis in systemic sclerosis. J Dermatol. 2010. 37:11–25.

11. Bhogal RK, Stoica CM, McGaha TL, Bona CA. Molecular aspects of regulation of collagen gene expression in fibrosis. J Clin Immunol. 2005. 25:592–603.

12. Chen SJ, Yuan W, Mori Y, Levenson A, Trojanowska M, Varga J. Stimulation of type I collagen transcription in human skin fibroblasts by TGF-beta: involvement of Smad 3. J Invest Dermatol. 1999. 112:49–57.

13. Ghosh AK. Factors involved in the regulation of type I collagen gene expression: implication in fibrosis. Exp Biol Med (Maywood). 2002. 227:301–314.

14. Sephel GC, Davidson JM. Elastin production in human skin fibroblast cultures and its decline with age. J Invest Dermatol. 1986. 86:279–285.

15. Neumann E, Gay RE, Gay S, Müller-Ladner U. Functional genomics of fibroblasts. Curr Opin Rheumatol. 2004. 16:238–245.

16. Wiedłocha A, Sørensen V. Signaling, internalization, and intracellular activity of fibroblast growth factor. Curr Top Microbiol Immunol. 2004. 286:45–79.

17. Chiquet M, Renedo AS, Huber F, Flück M. How do fibroblasts translate mechanical signals into changes in extracellular matrix production? Matrix Biol. 2003. 22:73–80.

18. Nagai Y, Miyata K, Sun GP, Rahman M, Kimura S, Miyatake A, et al. Aldosterone stimulates collagen gene expression and synthesis via activation of ERK1/2 in rat renal fibroblasts. Hypertension. 2005. 46:1039–1045.

19. Sato M, Shegogue D, Gore EA, Smith EA, McDermott PJ, Trojanowska M. Role of p38 MAPK in transforming growth factor beta stimulation of collagen production by scleroderma and healthy dermal fibroblasts. J Invest Dermatol. 2002. 118:704–711.