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Journal of Natural Medicines

, Volume 71, Issue 4, pp 723–734 | Cite as

Shikonin changes the lipopolysaccharide-induced expression of inflammation-related genes in macrophages

  • Lucia Satiko Yoshida
  • Tomohito Kakegawa
  • Yasukatsu Yuda
  • Hiromi Takano-Ohmuro
Original Paper
  • 303 Downloads

Abstract

We aimed to find candidate molecules possibly involved in the anti-inflammatory activity of shikonin (active compound of “Shikon”) by analyzing its effects on gene expression of lipopolysaccharide (LPS)-treated THP-1 macrophages. Polysome-associated mRNAs (those expected to be under translation: translatome) from cells treated with LPS alone (LPS: 5 µg/mL), shikonin alone (S: 100 nM), or LPS plus shikonin (LPS&S) for 3 h were analyzed by DNA microarray followed by detection of enriched pathways/gene ontologies using the tools of the STRING database. Candidate genes in enriched pathways in the comparison of LPS&S cells vs. LPS cells were analyzed by reverse-transcription quantitative real-time PCR (RT-qPCR; 1, 2, and 3 h). DNA microarray showed shikonin significantly influences gene expression. Gene expression changes between LPS&S cells and LPS cells were compared to detect relevant proteins and/or mRNAs underlying its anti-inflammatory effects: shikonin downregulated pathways which were upregulated in LPS cells, for example, ‘innate immune response’. Within changed pathways, three genes were selected for RT-qPCR analyses as key candidates influencing inflammatory responses: CYBA (component of the superoxide-generating Nox2 enzyme), GSK3B (controller of cell responses after toll-like receptor stimulation), and EIF4E (a key factor of the eukaryotic translation initiation factor 4F complex that regulates abundance of other proteins involved in immune functions). All three mRNAs were decreased at 2 h, and CYBA continued low at 3 h relative to LPS cells. Given that shikonin decreased the expression of CYBA gene of Nox2, in addition to the direct inhibition of the Nox2 activity that we have previously shown, it is suggested that one of its anti-inflammatory mechanisms could be attenuation of oxidative stress.

Keywords

Shikonin Naphthoquinone Translatome DNA microarray THP-1 macrophage Gene ontology 

Abbreviations

DMSO

Dimethylsulfoxide

FDR

False discovery rate

GO

Gene ontology

IL

Interleukin

LPS

Lipopolysaccharide

LPS cells

Cells treated with LPS alone

LPS&S cells

Cells treated with LPS and shikonin

S cells

Cells treated with shikonin alone

MAPK

Mitogen-activated protein kinases

NF-κB

Nuclear factor kappa B

NOS

Nitric oxide synthase

PPI

Protein–protein interaction

PMA

Phorbol myristate acetate

RT-qPCR

Reverse-transcription quantitative real-time PCR

ROS

Reactive oxygen species

TNF

Tumor necrosis factor

Notes

Acknowledgements

This research was funded by “GAKUIN TOKUBETSU KENKYUHI” grant for academic research (Musashino University) to H. Takano-Ohmuro.

Compliance with ethical standards

Conflict of interest

The authors have nothing to declare.

Supplementary material

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Copyright information

© The Japanese Society of Pharmacognosy and Springer Japan KK 2017

Authors and Affiliations

  • Lucia Satiko Yoshida
    • 1
  • Tomohito Kakegawa
    • 2
  • Yasukatsu Yuda
    • 1
  • Hiromi Takano-Ohmuro
    • 1
  1. 1.Research Institute of Pharmaceutical SciencesMusashino UniversityNishitokyoJapan
  2. 2.Faculty of Pharmaceutical SciencesJosai International UniversityToganeJapan

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