Interaction of Adipokines and Hepatitis B Virus on Histological Liver Injury in the Chinese

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1 132 ORIGINAL CONTRIBUTIONS nature publishing group Interaction of Adipokines and Hepatitis B Virus on Histological Liver Injury in the Chinese Vincent Wai-Sun Wong, MD1, 2, Grace Lai-Hung Wong, MD 1, 2, Ju n Yu, PhD 1, 2, Pau l C he u ng - Lu ng C hoi, FRCPath3, Anthony Wing-Hung Chan, MRCPath 3, Hoi - Yu n C han, MSc1, 2, E ag l e Siu - Hong C hu, MSc 1, 2, Alfred Sze-Lok Cheng, PhD 1, 2, Angel Mei-Ling Chim, MSc1, 2, Franc is Ka - L e u ng C han, MD 1, 2, Joseph Jao-Yiu Sung, MD, PhD 1, 2 and Henry Lik-Yuen Chan, MD1, 2 OBJECTIVES: METHODS: Chronic hepatitis B patients with diabetes and metabolic syndrome are at increased risk of cirrhosis and hepatocellular carcinoma, but the underlying mechanism is unclear. Our objective was to test whether dysregulation of adipokines contributes to liver injury. We also studied whether viral factors affected adipokines, insulin resistance, and hepatic steatosis. A prospective cohort of 266 chronic hepatitis B patients undergoing liver biopsy was studied. Fasting blood was taken for the analysis of tumor necrosis factor- α (TNF- α ), interleukin-6 (IL-6), adiponectin, leptin, and resistin. Insulin resistance was assessed by the homeostasis model assessment of insulin resistance (HOMA-IR). Factors associated with significant necroinflammation and cirrhosis were identified. RESULTS: Histological activity index was correlated with serum TNF- α ( R = 0.40, P < ) and IL-6 ( R = 0.32, P < ) but not with adiponectin, leptin, or resistin. By multivariate analysis, TNF- α was associated with significant necroinflammation after adjusting for age and viral factors (odds ratio (OR) 1.041, 95 % confidence interval (CI) , P = 0.04). Serum adiponectin had positive correlation with hepatitis B virus DNA ( R = 0.17, P = 0.007) and was decreased in patients with insulin resistance and hepatic steatosis. On the other hand, viral load, hepatitis B e-antigen status, and genotypes had no association with insulin resistance, hepatic steatosis, and the levels of TNF- α and IL-6. A total of 68 (25.6 % ) patients had cirrhosis. HOMA-IR, but not adipokine dysregulation, was independently associated with cirrhosis (OR 1.09, 95 % CI , P = 0.006). CONCLUSIONS: TNF- α and / or IL-6 contribute to hepatic necroinflammation in chronic hepatitis B patients. Adiponectin protects against insulin resistance and hepatic steatosis but does not affect liver injury. Adipokines and viral factors contribute to liver injury independently. Am J Gastroenterol 2010; 105: ; doi: /ajg ; published online 6 October 2009 INTRODUCTION Chronic hepatitis B affects more than 350 million people worldwide, and is one of the leading causes of liver cirrhosis and hepatocellular carcinoma (1). Disease progression has been shown to be associated with host factors (age, gender, and immune status), viral factors (hepatitis B viral load, genotype, basal core promoter, and other mutations), lifestyle (alcohol and smoking), and viral superinfection (hepatitis C, hepatitis D, and human immunodeficiency virus) (2). There is a growing epidemic of obesity and metabolic syndrome in both developed and developing countries. Recent data suggest that metabolic factors may also affect the natural history of chronic hepatitis B. Chronic hepatitis B patients with metabolic syndrome have higher risk of advanced fibrosis and cirrhosis (3). Diabetes and obesity are also independently associated with hepatocellular carcinoma development in patients with chronic viral hepatitis (4,5). The mechanism of how metabolic factors influence disease progression is not completely understood. 1 Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong, China ; 2 Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong, China ; 3 Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Hong Kong, China. Correspondence: Henry Lik-Yuen Chan, MD, Department of Medicine and Therapeutics, The Chinese University of Hong Kong, 9 / F, Prince of Wales Hospital, Ngan Shing Road, Hong Kong, Shatin, China. hlychan@cuhk.edu.hk Received 20 June 2009; accepted 1 September 2009

2 Adipokines and Chronic Hepatitis B 133 In patients with nonalcoholic fatty liver disease (NAFLD), dysregulated production of adipokines is a prominent feature (6,7). Adipose tissue secretes a number of bioactive proteins collectively known as adipokines, including leptin, adiponectin, resistin, tumor necrosis factor- α (TNF-α ), and interleukin- 6 (IL-6). They are bioactive proteins with effect on insulin resistance and inflammation. Adiponectin improves insulin sensitivity, reduces inflammation, and suppresses TNF- α production (8). Intrahepatic production of TNF- α is increased in NAFLD patients, especially among those with liver fibrosis (9). As hepatic steatosis is common in both the general population and chronic hepatitis B patients (10,11), the role of adipokines in chronic hepatitis B warrants further investigation. In this study, we investigated whether dysregulation of adipokines contributes to liver injury in a prospective cohort of chronic hepatitis B patients. We also studied whether viral factors affect adipokines, insulin resistance, and hepatic steatosis. Education Program with modifications by the International Diabetes Federation (12). Laboratory tests On the day of liver biopsy, a venous blood sample was taken after 8-h fasting for liver function tests, fasting glucose, lipids, and high-density lipoprotein and low-density lipoprotein cholesterol. Hepatitis B e-antigen (HBeAg) and antibody against HBeAg were measured by enzyme-linked immunosorbent assay (Sanofi Diagnostics, Pasteru, France). HBV DNA was measured by TaqMan real-time polymerase chain reaction using Eurohep standard to quantify the viral load (13). The range of HBV DNA detection is from 10 2 to 10 9 DNA copies per ml. HBV genotypes were determined by restriction fragment length polymorphism, as described previously (14). Leptin, adiponectin, TNF- α, IL-6, and resistin were measured by Bio-Plex Pro Human Diabetes Panel (Bio-Rad, Hercules, CA). Insulin was measured by enzymelinked immunosorbent assay (Dako, Glostrup, Denmark). METHODS Patients From July 2005 to October 2008, 266 consecutive patients with chronic hepatitis B undergoing liver biopsy were prospectively recruited from the hepatology clinic, Prince of Wales Hospital. The indications for liver biopsy included pretreatment evaluation and disease staging. The inclusion criteria included age 18 years or older, positive hepatitis B surface antigen for at least 6 months, and provision of informed written consent. We excluded patients with hepatitis C virus or human immunodeficiency virus coinfection, significant alcohol consumption (more than 30 g per week in men and 20 g per week in women), active malignancy, and contraindications to liver biopsy such as bleeding tendency and ascites. As antiviral treatment, in particular interferon, may affect both hepatitis B virus (HBV) and metabolic profile, only treatment-naive patients were included in this study. Similarly, we excluded patients on oral hypoglycemic agents or insulin therapy because of potential interference with adipokines and insulin resistance assessment. The study protocol was approved by the Clinical Research Ethics Committee of the Chinese University of Hong Kong. Clinical evaluation On the day of liver biopsy, body height, body weight, waist circumference, hip circumference, and blood pressure were measured. Body height and weight were measured using the same scale. Waist circumference was measured with a tape ruler at a level midway between the lower rib margin and iliac crest with the tape all around the body in the horizontal position. Central obesity was defined as waist circumference 90 cm in men and 80 cm in women (12). Body mass index (BMI) was calculated as body weight (kg) divided by height (m) squared. Obesity was defined as BMI 25 kg /m2 according to Asian cutoffs. Metabolic syndrome was defined according to the Adult Treatment Panel III of the National Cholesterol Liver histology Liver biopsy was performed using a 16G Temno needle (Care- Fusion, San Diego, CA). Histological scoring was performed by two pathologists (P.C.L.C. and A.W.H.C.) who were blinded to all clinical data except the diagnosis of chronic hepatitis B. Necroinflammation was scored by the modified Knodell scoring system (0 18) and fibrosis stage was scored by the Ishak scoring system (0 6) (15). Significant necroinflammation was defined as histological activity index 5 or above. Stage 5 or 6 fibrosis was considered as cirrhosis. Steatosis was assessed as the percentage of hepatocytes containing macrovesicular fat droplets, with grade 0 = < 5 % of hepatocytes affected, 1 = 5 % < 33 % of hepatocytes affected, 2 = % of hepatocytes affected, and 3 = >66 % of hepatocytes affected. Data analysis The correlation of serum adipokine levels with histological necroinflammation and fibrosis was tested by Spearman s test. The correlation between serum adipokine levels and HBV DNA was tested by Pearson s test. HBV DNA was logarithmically transformed for analysis. Serum adipokine levels in patients with positive and negative HBeAg and different genotypes were compared using the Mann Whitney U-test. The binary logistic regression model was performed to identify factors independently associated with significant necroinflammation and cirrhosis. Factors with P -values below 0.05 in the univariate analysis were included in the multivariate regression analysis. All analyses were performed using the Statistical Package for Social Sciences (SPSS, version 16.0; Chicago, IL). A two-sided P value of < 0.05 was considered statistically significant. RESULTS The clinical characteristics of 266 chronic hepatitis B patients were shown in Table 1. The mean age was 45 ± 11 years, and by the American College of Gastroenterology

3 134 Wong et al. Table 1. Clinical characteristics of 266 chronic hepatitis B patients Characteristic All ( N =266) Significant necroinflammation ( n =126) No significant necroinflammation ( n =140) P value Cirrhosis ( n = 68) No cirrhosis ( n =198) P value Age (years) 45 ± ± ± ± 9 43 ± Male gender, n ( % ) 203 (76.3) 96 (76.2) 107 (76.4) (77.9) 150 (75.8) 0.72 Hypertension, n ( % ) 29 (10.9) 10 (7.9) 19 (13.6) (14.7) 19 (9.6) 0.24 Diabetes, n ( % ) 22 (8.3) 8 (6.3) 14 (10.0) (14.7) 12 (6.1) Metabolic syndrome, n ( % ) 38 (14.3) 19 (15.1) 19 (13.6) (23.5) 22 (11.1) BMI (kg / m 2 ) 24.3 ± ± ± ± ± Male 24.6 ± ± ± ± ± Female 23.5 ± ± ± ± ± Waist circumference (cm) 84.3 ± ± ± ± ± Male 86.2 ± ± ± ± ± Female 77.5 ± ± ± ± ± HBV DNA (log copies per ml) 6.55 ± ± ± 1.84 < ± ± Positive HBeAg 114 (43.2) 68 (54.4) 46 (33.1) (44.1) 86 (43.4) 0.92 Genotypes, n ( % ) B 88 (33.1) 39 (31.0) 49 (35.0) 23 (33.8) 65 (32.8) C 161 (60.5) 85 (67.5) 76 (54.3) 43 (63.2) 118 (59.6) Unknown 17 (6.4) 2 (1.6) 15 (10.7) 2 (2.9) 15 (7.6) ALT (IU / l) 73 (60 93) 114 (79 150) 51 (34 73) < (46 100) 77 (47 126) 0.14 Total cholesterol HDL cholesterol LDL cholesterol 5.3 ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± Triglycerides 1.0 ( ) 1.0 ( ) 1.0 ( ) ( ) 1.0 ( ) Fasting glucose 5.5 ± ± ± ± ± Insulin (pmol / l) 60 (33 188) 67 (38 209) 57 (28 161) (37 468) 57 (30 154) HOMA-IR ( % ) 1.1 ( ) 1.2 ( ) 1.1 ( ) ( ) 1.1 ( ) TNF- α (pg / ml) 8.0 ( ) 8.9 ( ) 6.8 ( ) < ( ) 8.2 ( ) Adiponectin ( μ g / ml) 19 (12 30) 18 (12 36) 20 (13 29) (12 32) 20 (13 30) 0.61 IL-6 (pg / ml) 9.0 ( ) 9.4 ( ) 7.9 ( ) < ( ) 9.1 ( ) 0.21 Leptin (ng / ml) 2.3 ( ) 2.3 ( ) 2.3 ( ) ( ) 2.3 ( ) 0.70 Resistin (ng / ml) 1.9 ( ) 1.9 ( ) 1.9 ( ) ( ) 2.0 ( ) Steatosis grade 0 (0 1) 0 (0 1) 0 (0 1) (0 1) 0 (0 1) 0.20 Histological activity index 4 (2 5) 5 (5 7) 2 (2 3) < (2 6) 5 (3 5) Fibrosis stage 3 (2 5) 3 (2 4) 4 (2 5) (5 6) 3 (1 3) < ALT, alanine aminotransferase; BMI, body mass index; HBeAg, hepatitis B e-antigen; HBV, hepatitis B virus; HDL, high-density lipoprotein; HOMA-IR, homeostasis model assessment of insulin resistance; IL-6, interleukin-6; LDL, low-density lipoprotein; TNF- α, tumor necrosis factor- α. Numbers given in brackets are interquartile ranges.

4 Adipokines and Chronic Hepatitis B 135 (76.3 % ) were males. A total of 62 (30.5 % ) men and 20 (31.7 % ) women had central obesity. In total, 29 (10.9 % ) patients suffered from hypertension, 22 (8.3 % ) suffered from diabetes, and 38 (14.3 % ) had metabolic syndrome. Hepatic steatosis was present in 116 (43.6 % ) patients: grade 1 in 95 (35.7 % ), grade 2 in 17 (6.4 % ), and grade 3 in 4 (1.5 % ). The median histological activity index was 4 (interquartile range, 2 5). A total of 68 (25.6 % ) patients had cirrhosis. No patient had hepatic decompensation. HBV, adipokines, and steatosis Among 116 patients with hepatic steatosis, 58 (50.0 % ) were obese, 53 (45.7 % ) had central obesity, and 31 (26.7 % ) had metabolic syndrome. In contrast, among 150 patients without steatosis, 27 (18.0 % ) were obese, 29 (19.3 % ) had central obesity, and 7 (4.7 % ) had metabolic syndrome ( P < for all comparisons with patients with steatosis). The BMI was 26.1 ±4.9 kg /m2 in patients with steatosis and 22.9 ±2.8 kg /m 2 in those without steatosis ( P < ). Similarly, patients with steatosis had higher waist circumference (88.8 ±11.7 cm vs ±8.2 cm; P < ). BMI had negative correlation with serum adiponectin ( R = 0.24, P < ) and positive correlation with HOMA- IR ( R = 0.41, P < ). On the other hand, BMI had no significant correlation with TNF- α (R = 0.042, P = 0.51), IL-6 ( R = 0.023, P = 0.72), leptin ( R = 0.11, P = 0.088), and resistin ( R = 0.063, P = 0.33). Similarly, waist circumference had negative correlation with adiponectin ( R = 0.30, P < ) and positive correlation with HOMA-IR ( R = 0.33, P < ). There was no significant correlation between waist circumference and TNF- α ( R = 0.069, P = 0.29), IL-6 ( R = 0.042, P = 0.52), leptin ( R = 0.064, P = 0.33), and resistin ( R = 0.037, P = 0.57). HBV DNA had positive correlation with serum adiponectin ( R = 0.17, P = 0.007), but not with TNF- α ( R = 0.11, P = 0.074), IL-6 ( R = 0.090, P = 0.14), leptin ( R = 0.006, P = 0.93), resistin ( R = 0.054, P = 0.38), or HOMA-IR ( R = 0.059, P = 0.34). There was no significant difference in all adipokine levels and HOMA-IR between patients with positive and negative HBeAg and patients with genotype B and C virus HBV infection ( Table 2 ). These patients also had similar BMI and waist circumference. Insulin resistance, as measured by HOMA-IR, had negative correlation with serum adiponectin ( R = 0.19, P = 0.002), but positive correlation with leptin ( R = 0.26, P < ) and resistin ( R = 0.14, P = 0.019). Steatosis grade was inversely correlated with serum adiponectin, but positively correlated with IL-6, leptin, and HOMA-IR ( Table 3 ). Table 2. Metabolic and adipokine profiles of chronic hepatitis B patients with positive and negative HbeAg and genotype B and C virus Variable HBeAg positive ( n =116) HBeAg negative ( n =150) P value Genotype B ( n =88) Genotype C ( n =161) P value Body mass index (kg / m 2 ) 24.4 ± ± ± ± Male 24.8 ± ± ± ± Female 23.3 ± ± ± ± Waist circumference (cm) 84.2 ± ± ± ± Male 86.8 ± ± ± ± Female 76.2 ± ± ± ± HOMA-IR (%) 1.2 ( ) 1.1 ( ) ( ) 1.3 ( ) 0.91 TNF- α (pg/ ml) 8.4 ( ) 7.6 ( ) ( ) 8.1 ( ) 0.97 Adiponectin ( μ g / ml) 19 (12 29) 19 (12 32) (12 34) 18 (13 29) 0.69 IL-6 (pg/ ml) 9.2 ( ) 8.6 ( ) ( ) 8.8 ( ) 0.42 Leptin (ng/ ml) 2.2 ( ) 2.4 ( ) ( ) 2.2 ( ) 0.33 Resistin (ng/ ml) 1.9 ( ) 1.9 ( ) ( ) 1.8 ( ) 0.17 Steatosis grade 0 (0 1) 0 (0 1) (0 1) 0 (0 1) 0.91 Histological activity index 5 (3 7) 3 (2 5) < (2 5) 5 (3 7) Fibrosis stage 3 (2 5) 3 (1 5) (1 5) 3 (3 5) HBeAg, hepatitis B e-antigen; HOMA-IR, homeostasis model assessment of insulin resistance; IL-6, interleukin-6; TNF- α, tumor necrosis factor- α by the American College of Gastroenterology

5 136 Wong et al. Table 3. Correlation between adipokines, insulin resistance, and histological features Adipokine Steatosis grade Histological activity index Fibrosis stage R P value R P value R P value TNF- α < Adiponectin 0.33 < IL < Leptin Resistin HOMA-IR 0.28 < HOMA-IR, homeostasis model assessment of insulin resistance; IL-6, interleukin-6; TNF- α, tumor necrosis factor- α. R = correlation coeffi cient. Th ere was no significant correlation between HBV DNA and steatosis grade ( R = 0.002, P = 0.98). HBV DNA was 6.59 ± 1.68 log copies per ml among subjects with steatosis and 6.51 ± 1.87 log copies per ml among subjects without steatosis ( P = 0.70). A total of 52 (45.2 % ) of 116 patients with steatosis and 62 (41.6 % ) of 150 patients without steatosis had positive HBeAg ( P = 0.59). In patients with steatosis, 38 (32.8 % ) had genotype B HBV and 72 (62.1 % ) had genotype C HBV. In patients without steatosis, 50 (33.3 % ) had genotype B HBV and 89 (59.3 % ) had genotype C HBV ( P = 0.75 compared with patients with steatosis). HBV, adipokines and necroinflammation A total of 126 patients (47.4 % ) had significant necroinflammation. TNF- α, IL-6, and leptin had moderate positive correlation with hepatic necroinflammation ( Table 3 ). On the other hand, necroinflammation was not correlated with adiponectin or resistin. As cirrhosis might have a direct effect on insulin resistance and adipokines, the test was repeated in noncirrhotic patients. Among patients with stage 0 4 fibrosis, TNF- α ( R = 0.39, P < ) and IL-6 ( R = 0.30, P < ) also had moderate correlation with necroinflammation. Patients with significant necroinflammation were younger and had higher HBV DNA and ALT levels ( Table 1 ). They were more likely to have positive HBeAg. Serum TNF- α and IL-6 levels were significantly higher in patients with significant necroinflammation. After adjusting for age, ALT, HBV DNA, and HBeAg status, serum TNF- α remained an independent factor associated with significant necroinflammation ( Table 4 ). In contrast, IL-6 was no longer associated with significant necroinflammation after adjusting for these factors (odds ratio 1.004; 95 % confidence interval ; P = 0.53). HBV, adipokines, and cirrhosis Patients with cirrhosis were older and more likely to have diabetes, obesity, and metabolic syndrome ( Table 1 ). They had significantly higher BMI, waist circumference, fasting insulin, Table 4. Multivariate analysis of factors associated with significant necroinflammation and cirrhosis Factor Odds ratio Significant necroinflammation 95 % confidence interval P value Age (years) ALT (IU / l) HBV DNA (log copies per ml) < Positive HBeAg TNF- α (ng / ml) Cirrhosis Age (years) HOMA-IR ( % ) Metabolic syndrome TNF- α (ng / ml) ALT, alanine aminotransferase; HBV, hepatitis B virus; HOMA-IR, homeostasis model assessment of insulin resistance; TNF- α, tumor necrosis factor- α. and HOMA-IR. On the other hand, serum TNF- α was lower in patients with cirrhosis. By Spearman s test, the fibrosis stage had a mild negative correlation with serum TNF- α and positive correlation with HOMA-IR. After adjusting for age and metabolic syndrome, HOMA-IR but not TNF- α remained associated with cirrhosis ( Table 4 ). DISCUSSION In this study, we tested the hypothesis that adipokines contribute to liver injury in chronic hepatitis B patients, thus explaining the observation that metabolic factors affect liver fibrosis and hepatocellular carcinoma development. Serum TNF- α and IL-6 were increased in patients with significant necroinflammation.

6 Adipokines and Chronic Hepatitis B 137 Serum adiponectin had positive correlation with HBV DNA, and was decreased in patients with insulin resistance and hepatic steatosis. On the other hand, viral load, HBeAg status, and genotypes had no association with insulin resistance and hepatic steatosis. TNF- α is a pro-inflammatory cytokine implicated in various liver diseases such as chronic hepatitis C, alcoholic liver disease, and NAFLD (7,16 20). Administration of pentoxifylline, a TNF- α inhibitor, may improve ALT levels in patients with alcoholic hepatitis and NAFLD (21,22). In this study, TNF- α was also independently associated with necroinflammation in chronic hepatitis B patients. In contrast, serum TNF- α was lower in patients with cirrhosis. This reflects that necroinflammatory activity tends to regress as fibrosis progresses. This phenomenon is supported by the fact that patients with cirrhosis in this study also tended to have lower histological activity index. Similarly, IL-6 was another pro-inflammatory cytokine that was increased in chronic hepatitis B patients with significant necroinflammation, although the association was not statistically significant after adjusting for age, ALT, HBV DNA, and HBeAg. In mice receiving partial hepatectomy, prolonged administration of IL-6 reduces survival (23). Recently, IL-6 was also implicated in hepatocarcinogenesis. In a murine model of hepatocellular carcinoma, mice produce high levels of IL-6 during liver injury (24). The development of hepatocellular carcinoma can be abolished by ablation of IL-6. In humans with chronic hepatitis B, high serum IL-6 level is also associated with future hepatocellular carcinoma development (25). Moreover, high IL-6 production is associated with diabetes (26). This further supports the observation that diabetes and insulin resistance are more common in chronic hepatitis B patients with cirrhosis, and that diabetes is independently associated with hepatocellular carcinoma development (4). Interestingly, HBV DNA was found to have a positive correlation with serum adiponectin. In turn, serum adiponectin was decreased in patients with insulin resistance and hepatic steatosis. Adiponectin is an important mediator of insulin resistance and is closely implicated in the pathogenesis of NAFLD (27). Adiponectin is lower in NAFLD subjects than in healthy controls (6,7,20). In the liver, adiponectin suppresses fatty acid synthesis and promotes mitochondrial β-oxidation. Adiponectin also opposes the synthesis and release of TNF- α from macrophages in adipose tissue, resulting in antiinflammatory action (28). Although the association between HBV DNA and adiponectin needs further confirmation, it has been shown in large Chinese cohorts that chronic hepatitis B patients have a lower prevalence of metabolic syndrome than does the general population (29,30). The effect is largely driven by lower serum triglycerides in chronic hepatitis B patients. Whether adiponectin and insulin resistance have separate effects warrants further investigations (31,32). On the other hand, adiponectin was not associated with hepatic necroinflammation and fibrosis. The association between adiponectin and liver injury in chronic hepatitis is controversial. In a study of 100 Chinese chronic hepatitis B patients, high serum adiponectin level was associated with advancing fibrosis stage (33). In contrast, adiponectin level was inversely correlated to ALT level in another study of 21 chronic hepatitis B patients (34). In those studies, increased adiponectin level might be secondary to active hepatic injury instead of directly contributing to fibrosis progression. Besides, the use of insulin sensitizers was not an exclusion criterion. As the prevalence of diabetes is high among cirrhotic patients, the use of insulin sensitizers might also affect adipokine levels. In contrast, our study excluded patients on insulin or oral hypoglycemic agents, and the findings were more in line with larger studies on chronic hepatitis C that showed an association between hypoadiponectinemia and steatosis but not fibrosis stage (35,36). Our study has a few limitations. First, patients on antidiabetic treatment were excluded. The possibility that adipokines, insulin resistance, and metabolic syndrome may have a stronger role in liver injury in patients with more severe diabetes cannot be excluded. However, our analysis avoided the potential interference of antidiabetic treatment on the assessment of adipokines and insulin resistance. Second, all subjects were ethnic Chinese. In NAFLD studies, an association between disease severity and different gene polymorphisms has been reported in different ethnic groups (20,37). Our findings need to be confirmed in other countries. In summary, chronic hepatitis B patients may have a lower risk of metabolic syndrome because of the increased production of adiponectin. Ongoing liver injury appears to be mediated through the increased production of TNF- α and /or IL-6. As viral factors are not associated with insulin resistance or pro-inflammatory adipokines, adipokines and the virus probably contribute to liver injury independently. CONFLICT OF INTEREST Guarantor of the article : Henry Lik-Yuen Chan, MD. Specific author contributions: Conception: Vincent W.S. Wong, Jun Yu, and Henry L.Y. Chan; study design: Vincent W.S. Wong, Grace L.H. Wong, Jun Yu, and Henry L.Y. Chan; drafting the article: Vincent W.S. Wong and Henry L.Y. Chan; data analysis, acquisition, and interpretation: Vincent W.S. Wong, Grace L.H. Wong, Angel M.L. Chim, and Henry L.Y. Chan; pathological assessment: Paul C.L. Choi and Anthony W.H. Chan; laboratory tests: Jun Yu, Hoi-Yun Chan, Eagle S.H. Chu, and Alfred S.L. Cheng. All the authors reviewed the manuscript and approved the final version. Financial support: The study was partially supported by the Research Fund of the Department of Medicine and Therapeutics, The Chinese University of Hong Kong. Potential competing interests: Joseph J.Y. Sung received lecture fees from AstraZeneca Hong Kong Limited, GSK Pharmaceuticals International, and Roche. Henry L.Y. Chan is a member of the advisory boards of Novartis Pharmaceutical, Schering-Plough Corporation, Bristol-Myers Squibb, and Pharmasset by the American College of Gastroenterology

7 138 Wong et al. Study Highlights WHAT IS CURRENT KNOWLEDGE 3 Chronic hepatitis B patients with diabetes and metabolic syndrome are at increased risk of cirrhosis and hepatocellular carcinoma, but the underlying mechanism is unclear. 3 Dysregulation of adipokines is associated with hepatic steatosis and liver injury in nonalcoholic fatty liver disease. It is uncertain whether they contribute to liver injury in chronic hepatitis B. WHAT IS NEW HERE 3 Higher hepatitis B virus DNA was associated with higher adiponectin, which might contribute to reduced hepatic steatosis. 3 Significant hepatic necroinflammation is associated with increased serum levels of tumor necrosis factor- α and interleukin-6. 3 Viral load and genotypes are not associated with abnormal levels of tumor necrosis factor- α and interleukin-6, suggesting that adipokines and the virus contribute to liver injury independently. REFERENCES 1. Chan HL, Sung JJ. Hepatocellular carcinoma and hepatitis B virus. Semin Liver Dis 2006 ;26 : L i aw Y F, S ol l ano J D. Fa c tors i n fluencing liver disease progression in chronic hepatitis B. Liver Int 2006 ; 26 : Wong G L, Wong V W, C hoi P C et al. Metabolic syndrome increases the risk of liver cirrhosis in chronic hepatitis B. Gut 2009 ; 58 : L ai M S, Hsi eh M S, C hiu Y H et al. Type 2 diabetes and hepatocellular carcinoma: a cohort study in high prevalence area of hepatitis virus infection. Hepatology 2006 ; 43 : Yu MW, Shih WL, Lin CL et al. Body-mass index and progression of hepatitis B: a population-based cohort study in men. J Clin Oncol 2008 ;26 : Hui J M, Ho d ge A, Far rel l G C et al. Beyond insulin resistance in NASH: TNF-alpha or adiponectin? Hepatology 2004 ; 40 : Wong V W, Hui AY, Ts ang S W et al. Metabolic and adipokine profile of Chinese patients with nonalcoholic fatty liver disease. Clin Gastroenterol Hepatol 2006 ; 4 : Xu A, Wang Y, Ke shaw H et al. The fat-derived hormone adiponectin alleviates alcoholic and nonalcoholic fatty liver diseases in mice. J Clin Invest 2003 ;112 : C re sp o J, C ayon A, Fer nand e z - Gi l P et al. Gene expression of tumor necrosis factor alpha and TNF-receptors, p55 and p75, in nonalcoholic steatohepatitis patients. Hepatology 2001 ; 34 : Peng D, Han Y, D ing H et al. Hepatic steatosis in chronic hepatitis B patients is associated with metabolic factors more than viral factors. J Gastroenterol Hepatol 2008 ; 23 : Shi J P, Fan JG, Wu R et al. Prevalence and risk factors of hepatic steatosis and its impact on liver injury in Chinese patients with chronic hepatitis B infection. J Gastroenterol Hepatol 2008 ; 23 : A lb e r t i KG, Z im me t P, Shaw J. The metabolic syndrome--a new worldwide definition. Lancet 2005 ;366 : C han H L, C hu i A K, L au W Y et al. Factors associated with viral breakthrough in lamivudine monoprophylaxis of hepatitis B virus recurrence after liver transplantation. J Med Virol 2002 ;68 : C han H L, Ts ang S W, Wong M L et al. Genotype B hepatitis B virus is associated with severe icteric flare-up of chronic hepatitis B virus infection in Hong Kong. Am J Gastroenterol 2002 ;97 : Isha k K, B apt ist a A, Bi anch i L et al. Histological grading and staging of chronic hepatitis. J Hepatol 1995 ; 22 : Zy lb er b e rg H, R imani ol AC, Pol S et al. Soluble tumor necrosis factor receptors in chronic hepatitis C: a correlation with histological fibrosis and activity. J Hepatol 1999 ; 30 : C re sp o J, R ive ro M, Fabreg a E et al. Plasma leptin and TNF-alpha levels in chronic hepatitis C patients and their relationship to hepatic fibrosis. Dig Dis Sci 2002 ;47 : Maeno T, Okumura A, Ishikawa T et al. Mechanisms of increased insulin resistance in non-cirrhotic patients with chronic hepatitis C virus infection. J Gastroenterol Hepatol 2003 ; 18 : S ong Z, Z hou Z, Ur i ar te S et al. S-adenosylhomocysteine sensitizes to TNFalpha hepatotoxicity in mice and liver cells: a possible etiological factor in alcoholic liver disease. Hepatology 2004 ;40 : Wong V W, Wong G L, Ts ang S W et al. Genetic polymorphisms of adiponectin and tumor necrosis factor-alpha and nonalcoholic fatty liver disease in Chinese people. J Gastroenterol Hepatol 2008 ; 23 : A k r iv i a d is E, B ot l a R, Br i g g s W et al. Pentoxifylline improves short-term survival in severe acute alcoholic hepatitis: a double-blind, placebocontrolled trial. Gastroenterology 2000 ; 119 : S at ap at hy SK, G arg S, C hau han R et al. B ene fi cial effects of tumor necrosis factor-alpha inhibition by pentoxifylline on clinical, biochemical, and metabolic parameters of patients with nonalcoholic steatohepatitis. Am J Gastroenterol 2004 ;99 : Sun Z, Klein AS, Radaeva S et al. In vitro interleukin-6 treatment prevents mortality associated with fatty liver transplants in rats. Gastroenterology 2003 ;125 : Naugler WE, Sakurai T, Kim S et al. Gender disparity in liver cancer due to sex differences in MyD88-dependent IL-6 production. Science 2007 ;317 : Wong V W, Yu J, C he ng AS et al. High serum interleukin-6 level predicts future hepatocellular carcinoma development in patients with chronic hepatitis B. Int J Cancer 2009 ; 124 : L iu S, Tin ker L, S ong Y et al. A prospective study of inflammatory cytokines and diabetes mellitus in a multiethnic cohort of postmenopausal women. Arch Intern Med 2007 ;167 : Larter CZ, Farrell GC. Insulin resistance, adiponectin, cytokines in NASH: which is the best target to treat? J Hepatol 2006 ; 44 : Mas a k i T, C hib a S, Tat su k aw a H et al. Adiponectin protects LPS-induced liver injury through modulation of TNF-alpha in KK-Ay obese mice. Hepatology 2004 ;40 : Jan C F, C he n C J, C hiu Y H et al. A population-based study investigating the association between metabolic syndrome and hepatitis B/C infection (Keelung Community-based Integrated Screening study No. 10). Int J Obes (Lond) 2006 ;30 : Luo B, Wang Y, Wang K. Association of metabolic syndrome and hepatitis B infection in a Chinese population. Clin Chim Acta 2007 ;380 : Kumar M, C hou d hu r y A, Mang l i k N et al. Insulin resistance in chronic hepatitis B virus infection. Am J Gastroenterol 2009 ; 104 : Park SH, Kim DJ, Lee HY. Insulin resistance is not associated with histologic severity in nondiabetic, noncirrhotic patients with chronic hepatitis B virus infection. Am J Gastroenterol 2009 ; 104 : Hu i C K, Z hang H Y, L e e N P et al. Serum adiponectin is increased in advancing liver fibrosis and declines with reduction in fibrosis in chronic hepatitis B. J Hepatol 2007 ; 47 : Lu J Y, Su TC, L iu Y H et al. Lower plasma adiponectin is correlated to higher alanine aminotransferase independent of metabolic factors and hepatitis B virus carrier status. Intern Med J 2007 ; 37 : Cu a I H, Hu i J M, B and ar a P et al. Insulin resistance and liver injury in hepatitis C is not associated with virus-specific changes in adipocytokines. Hepatology 2007 ; 46 : Jonss on J R, Mos che n A R, Hick man I J et al. Adiponectin and its receptors in patients with chronic hepatitis C. J Hepatol 2005 ; 43 : Valenti L, Fracanzani AL, Dongiovanni P et al. Tumor necrosis factor alpha promoter polymorphisms and insulin resistance in nonalcoholic fatty liver disease. Gastroenterology 2002 ;122 :