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BioMed Research International
Volume 2013 (2013), Article ID 963457, 8 pages
Research Article

Revealing the Mechanism of In Vitro Wound Healing Properties of Citrus tamurana Extract

1Department of Applied Physiology, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan
2Department of Obstetrics and Gynecology, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan

Received 9 October 2012; Revised 26 February 2013; Accepted 28 March 2013

Academic Editor: Jorge Berlanga Acosta

Copyright © 2013 Madhyastha Harishkumar et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


In the present investigation, we examined the effect of Hyuganatsu (Citrus tamurana) extract (HE) on skin fibroblast (TIG-119) proliferation and migration during in vitro wound healing. HE selectively inhibited proliferation of TIG-119 cells at higher concentration (>1.0 mg/mL); at lower concentrations (0.1, 0.25, 0.5, and 0.75 mg/mL), it exhibited linear and time-dependent cell proliferation. In vitro scratch wound healing studies showed that the HE also accelerated the migration of cells towards the wounded region. Cytometric analysis demonstrated that HE extract did not alter G1/0 and S phases of cell cycle in any concentration studied; however, G2/M phases of cell cycle were significantly ( ) accelerated at 0.75 mg/mL dose. RT-PCR and Western blotting analysis indicated that HE markedly overexpressed levels of Rac-1, Rho-A, and Cdc-42 mRNA and the respective proteins. Cyclin-dependent kinases (Cdk-1 and -2) gene expression activity was significantly ( ) increased, but protein content decreased during treatment with HE. The induction of Cdk-1 and -2 by HE was abolished by inhibitors, transcription (DRB), and translation (CHX), implying transcriptional regulation that required de novo protein synthesis.