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. 2008 May 6;13(3):475. doi: 10.2478/s11658-008-0016-7

Hydrogen peroxide as a potential mediator of the transcriptional regulation of heparan sulphate biosynthesis in keratinocytes

Fumiaki Nakayama 1,✉, Akiko Hagiwara 1,2, Tetsuo Yamamoto 1,3, Makoto Akashi 1
PMCID: PMC6275676  PMID: 18463796

Abstract

Ionizing radiation is one of the types of oxidative stress that has a number of damaging effects on cutaneous tissues. One of the histological features of radiation-induced cutaneous fibrosis is the accumulation of extracellular matrix (ECM) components, including heparan sulfate proteoglycan (HSPG), which are required for the repair of tissue damage, and operate by interacting with a variety of growth factors. In this study, we established a model of human HaCaT keratinocytes overexpressing anti-oxidative enzyme genes to elucidate the mechanism of oxidative stress leading to the accumulation of HSPG and the role of its accumulation. Catalase overexpression induced an increase in anti-HS antibody (10E4) epitope expression in these cells. Western blotting showed that the smeared bands of HSPG were obviously shifted to a higher molecular weight in the catalase transfectants due to glycosylation. After heparitinase I treatment, the core proteins of HSPG were expressed in the catalase transfectants to almost the same extent as in the control cells. In addition, the transcript levels of all the enzymes required for the synthesis of the heparan sulfate chain were estimated in the catalase transfectant clones. The levels of five enzyme transcripts — xylosyltransferase-II (XT-II), EXTL2, D-glucuronyl C5-epimerase (GLCE), HS2-O-sulfotransferase (HS2ST), and HS6-O-sulfotransferase (HS6ST) — were significantly increased in the transfectants. Moreover, hydrogen peroxide was found to down-regulate the levels of these enzymes. By contrast, siRNA-mediated repression of catalase decreased 10E4 epitope expression, the transcript level of HS2ST1, and the growth rate of HaCaT cells. These findings suggested that peroxide-mediated transcriptional regulation of HS metabolism-related genes modified the HS chains in the HaCaT keratinocytes.

Key words: Heparan sulfate proteoglycan, HaCaT keratinocyte, Glycosyltransferase, Catalase, Sulfotransferase

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Abbreviations used

CS

chondroitin sulfate

FGF

fibroblast growth factor

GAG

glycosaminoglycan

Gal

galactose

GalNAc

N-acetylgalactosamine

GlcA

glucuronic acid

GLCE

d-glucuronyl C5-epimerase

GlcNAc

N-acetylglucosamine

HSPG

heparan sulfate proteoglycan

HS2ST

heparan sulfate 2-O-sulfotransferase

HS6ST

heparan sulfate 6-O-sulfotransferase

PDGF

platelet-derived growth factor

ROS

reactive oxygen species

SOD

superoxide dismutase

XT

xylosyltransferase

Xyl

xylose

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