Hepatol Res. 2013 Oct;43(10):1078-92.

Serum oxidative-anti-oxidative stress balance is dysregulated in patients with hepatitis C virus-related hepatocellular carcinoma.

Mamoru Nishimura1)2), Akinobu Takaki1), Naofumi Tamaki3), Takayuki Maruyama4), Hideki Onishi1), Sayo Kobayashi1), Kazuhiro Nouso1), Tetsuya Yasunaka1), Kazuko Koike1), Hiroaki Hagihara1), Kenji Kuwaki1), Shinichiro Nakamura1), Fusao Ikeda1), Yoshiaki Iwasaki5), Takaaki Tomofuji4), Manabu Morita4), Kazuhide Yamamoto1)

1)Department of Gastroenterology and Hepatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama 700-8558, Japan

2)Department of Gastroenterology, Okayama City Hospital, 6-10 Amase, Kita-ku, Okayama 700-8857, Japan

3)Department of Preventive Dentistry, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto-cho, Tokushima 770-8504, Japan

4)Department of Preventive Dentistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama 700-8558, Japan

5)Health Service Center, Okayama University, Okayama, Japan

1-1-1 Tsushima Naka, Kita-ku, Okayama 700-8530, Japan


Address for correspondence: Akinobu Takaki

2-5-1 Shikata-cho, Kita-ku, Okayama 700-8558, Japan

Fax: +81 86 225 5991; Tel: +81 86 235 7219

E-mail: akitaka@md.okayama-u.ac.jp

Running title: Oxidative stress balance in chronic hepatitis C



Aim: Oxidative stress is associated with progression of chronic liver disease (CLD). This association is best established in chronic hepatitis C. However, the anti-oxidative state is not well characterized. The objective of the present study was to investigate the balance of oxidative and anti-oxidative stress in CLD patients.

Methods: We recruited a study population of 208 patients, including healthy volunteers (HV; n = 15), patients with hepatitis B virus (HBV)-related CLD without or with hepatocellular carcinoma (HBV-nonHCC, n = 25, and HBV-HCC, n = 50, respectively), and patients with hepatitis C virus (HCV)-related CLD without or with HCC (HCV-nonHCC, n = 49, and HCV-HCC, n = 69, respectively). Serum levels of reactive oxygen metabolites (ROM) and anti-oxidative markers (OXY-adsorbent test; OXY) were determined, and the balance of these values was used as the oxidative index. Correlations among ROM, OXY, oxidative index, and clinical characteristics were investigated.

Results: Patients with CLD exhibited elevated ROM and oxidative index compared to HV. Among patients with CLD, HCV-positive status correlated with increased ROM. In CLD, HCV-HCC patients exhibited the highest ROM levels. Among HCV-related CLD patients, lower OXY correlated with HCC-positive status, but was recovered by eradication of HCC. In HCV-HCC, lower OXY correlated with high PT-INR.

Conclusions: HCV-positive CLD patients displayed higher oxidative stress and HCV-HCC patients displayed lower anti-oxidative state. Anti-oxidative state depression was associated with liver reservoir-related data in HCV-HCC and could be reversed with HCC eradication.



Oxidative stress is increasingly recognized as a key factor in the progression of chronic liver disease (CLD). Chronic hepatitis C has been noted as a source of strong oxidative stress, resulting in a high rate of hepatocarcinogenesis among patients with cirrhosis. The liver, a metabolically important organ, is a major reservoir of the mitochondria that serves as the source of reactive oxygen species (ROS). As a result, CLD could represent a major inducer of oxidative stress. In addition, HCV itself induces oxidative stress on hepatocytes. However, oxidative stress is physiologically important to protect cells from viral infection. ROS can rapidly inhibit HCV-RNA replication in HCV replicon-harboring cells, and antioxidants reverse oxidative effects and thereby permit HCV replication. Clinical studies of antioxidant therapies for chronic hepatitis C have yielded disappointing results.

The objective of the present study was to investigate the balance between oxidative and anti-oxidative stresses in patients with chronic viral liver diseases. Serum levels of reactive oxygen metabolites (ROM) were determined as a marker of circulating ROS. The OXY-adsorbent test was also performed to evaluate the corresponding anti-oxidative status.

Oxidative stress-related markers in CLD patients

Both the oxidative stress marker ROM and calculated oxidative index were significantly higher in patients with CLD than in the HV group (Fig. 1A). Among CLD patients, ROM differed significantly for HCV-nonHCC and HCV-HCC vs. HBV-HCC, OXY differed significantly for HCV-HCC vs. HBV-HCC and HCV-nonHCC, oxidative index differed significantly for HCV-HCC vs. all others (Fig. 1B).

In HCV-HCC, OXY correlated with PT-INR levels.

Follow-up of oxidative stress markers in HCV-positive CLD patients

Follow-up data after about one and a half years (median: 16 months for HCV-nonHCC; 17 months for HCV-HCC) were collected from HCV-positive CLD patients (Fig. 2A). In HCV-nonHCC, the three oxidative stress-related markers showed no change during follow-up. In HCV-HCC, OXY was increased in the complete remission (CR) group (Fig. 2B,C).


Study importance

In the present study, serum ROM was higher in patients with CLD especially in HCV positive patients. In contrast, anti-oxidative status OXY was attenuated in HCV-positive HCC patients. The follow-up data revealed that OXY attenuation in HCV-HCC could be reversed by eradication of HCC.

HCV-induced oxidative stress is thus implicated in the disease progression of chronic hepatitis.

Total serum antioxidant levels, as assessed by OXY, were diminished specifically in patients with HCV-related HCC. In HCV-positive HCC patients, higher PT-INR level was the only factor that correlated with lower antioxidant capacity. Liver reservoir function is involved in anti-oxidative state.

Indeed, our results revealed that oxidative index was independently elevated in HCV-positive and HCC-positive CLD patients. The balance of oxidative stress is more oxidative in HCV-positive HCC patients, and antioxidant therapy might be effective for such patients.

The follow-up study revealed that OXY reduction was recovered after CR of HCC. As the reduced anti-oxidative condition could be recovered by eradication of HCC, the anti-oxidative condition could be a result of HCV-positive HCC. To clarify whether oxidative stress is a cause or result of HCC, assessment of larger numbers of patients with longer follow-up is needed.

In conclusion, oxidative stress was higher in HCV-positive patients compared to healthy controls and HBV-positive patients with HCC. Anti-oxidative stress was lowest in HCV-positive HCC patients and could be recovered by tumor eradication. This distinction was most apparent in patients with higher PT-INR, suggesting that liver reservoir function may be important for maintaining anti-oxidative function. The oxidative-anti-oxidative balance was most biased towards the oxidative state in HCV-positive HCC patients.

20140323Fig1OXYBiomedFigureTakakiFigure 1: The distribution of ROM, OXY, and oxidative index in patient groups. (A) Comparison between HV and CLD: The ROM and oxidative index were higher in CLD patients. (B) Comparison among HBV-nonHCC, HBV-HCC, HCV-nonHCC, and HCV-HCC: Among the CLD groups, the ROM value was significantly higher (P < 0.05) in HCV-nonHCC and HCV-HCC compared to HBV-HCC. OXY was significantly lower (P < 0.05) in HCV-HCC compared to that in HBV-HCC and HCV-nonHCC. Oxidative index was significantly higher (P < 0.05) in HCV-HCC than in other CLD groups.

HV, healthy volunteers; CLD, chronic liver disease patients; HBV-nonHCC, hepatitis B virus (HBV)-related CLD without hepatocellular carcinoma (HCC); HBV-HCC, HBV-related CLD with HCC; HCV-nonHCC, HCV-related CLD without HCC; HCV-HCC, HCV-related CLD with HCC.


20140323Fig2OXYBiomedFigureTakakiFigure 2: Oxidative stress-related marker changes after about 1.5 years. (A) The follow-up data change in HCV-non HCC. No change was seen in this period (median interval, 16 months). (B) HCV-HCC with a treatment response of CR. OXY level increased after follow-up period (median, 17 months interval). (C) HCV-HCC with treatment response PD+SD+PR. No statistical differences were observed.

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