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Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Backgound:

Cyclooxygenase-2 may play a role in the development of hepatocellular carcinoma, but the relationship between cyclooxygenase-2 and chronic hepatitis B is unknown.

Aim:

To investigate the expression and cellular localization of cyclooxygenase-2 in chronic hepatitis B patients and the effects of anti-viral therapy.

Methods:

Using immunohistochemistry, in situ hybridization, Western blot and reverse transcription polymerase chain reaction, protein and messenger RNA expression and cellular localization of cyclooxygenase-2 in 35 chronic hepatitis B patients were assessed. Fourteen histologically normal and non-viral-infected livers were used as controls. The cyclooxygenase-2 immunoreactivities of paired liver biopsies from 12 patients receiving anti-viral therapy were compared.

Results:

Immunohistochemistry and in situ hybridization revealed that cyclooxygenase-2 expression was confined to hepatocytes. Patients with chronic hepatitis B had significantly higher cyclooxygenase-2 expression compared with controls. The cyclooxygenase-2 expression of hepatitis B e antigen-positive and -negative chronic hepatitis B patients was not significantly different, although the necro-inflammatory activity of the latter group was significantly lower. Over-expression of cyclooxygenase-2 in patients with chronic hepatitis B was further confirmed by Western blot and reverse transcription polymerase chain reaction. Twelve hepatitis B e antigen-positive chronic hepatitis B patients received anti-viral therapy: lamivudine in seven and interferon in five. Despite hepatitis B e antigen seroconversion, disappearance of hepatitis B virus DNA in serum, normalization of liver enzymes and a significant reduction in necro-inflammatory activity in all 12 patients, no significant change in cyclooxygenase-2 expression was found.

Conclusions:

Chronic hepatitis B is associated with elevated cyclooxygenase-2 levels in hepatocytes, and the over-expression of this enzyme does not reflect inflammatory activity. Up-regulation of cyclooxygenase-2 persists after successful anti-viral therapy.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Cyclooxygenase (COX) catalyses the rate-limiting step of prostaglandin biosynthesis. There are two COX isozymes, COX-1 being the constitutive isozyme and COX-2 being the inducible isoform.1 Over-expression of COX-2 has been reported in many gastrointestinal tumours2–10 and premalignant lesions.2, 3, 5, 6, 9–11 Previous studies have shown that COX-2 interferes with cellular adhesion, inhibits apoptosis of intestinal epithelial cells12 and increases cell proliferation.13, 14 COX-2-associated prostanoids are also mutagenic15, 16 and tumorigenic.17 Furthermore, COX-2 regulates angiogenesis,18, 19 enhances metastatic potential20 and inhibits immune surveillance,21 thus facilitating the growth and invasiveness of the tumour. All of these studies support the notion that COX-2 plays a key role in carcinogenesis.

Recently, up-regulation of COX-2 has been demonstrated in cirrhosis and well-differentiated hepatocellular carcinoma, but less so in poorly differentiated hepatocellular carcinoma, suggesting that COX-2 is involved in the early steps of hepatic carcinogenesis.5–7 Because previous studies have concentrated on hepatitis C virus (HCV)-associated chronic hepatitis, cirrhosis and hepatocellular carcinoma, the relationship between COX-2 and chronic hepatitis B remains unknown. On the other hand, there are strong suggestions that interferon treatment reduces the incidence of hepatocellular carcinoma in patients with HCV infection.22–24 However, the effect of interferon on the prevention of hepatocellular carcinoma in patients with hepatitis B virus (HBV) infection is less certain.25–27 In this study, we investigated the expression of COX-2 in chronic hepatitis caused by HBV, and followed the changes after anti-viral therapy with interferon or lamivudine. We hypothesized that HBV infection induces COX-2 expression in hepatocytes and that COX-2 up-regulation would be reversed after successful eradication of HBV.

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Patients and controls

Liver biopsy specimens from 35 patients with proven chronic hepatitis B infection were studied. All patients had elevation of serum alanine aminotransferase levels and detectable HBV DNA by bDNA assay (Chiron Diagnostic, Emeryville, CA, USA). Twenty-six patients had positive hepatitis B e antigen (HBeAg) and nine patients had negative HBeAg. Two donor livers, four histologically normal livers (non-tumour sites adjacent to liver metastasis), two with pyogenic cholangitis and six with non-alcoholic steato-hepatitis were used as controls. All control patients were negative for hepatitis B s antigen (HBsAg). The demographic, serological and histological data of the patients are shown in Table 1. Liver biopsy tissues were processed for routine histology, immunohistochemistry and in situ hybridization. The histology activity index28 of the specimens was scored by a single pathologist who had no knowledge of the COX-2 immunohistochemical staining results, and the necro-inflammatory score and fibrosis score were individually analysed. The remaining surgical specimens of HBV-infected and non-infected liver tissues were snap-frozen, stored at – 80 °C and used for Western blot and reverse transcription polymerase chain reaction.

Table 1.   Demographic, serological and histologual data pf the different patient groups Thumbnail image of

Twelve of the 26 HBeAg-positive chronic hepatitis B patients received anti-viral therapy: seven received lamivudine, 100 mg/day, for 1 year, and five received interferon, 5–10 MU, three times a week for 16–24 weeks. Follow-up liver biopsy specimens in the second year for patients on lamivudine and at 1 year post-treatment for patients on interferon were studied. All patients had HBeAg seroconversion, disappearance of HBV DNA in serum and normalization of liver enzymes at the time of follow-up. All pre- and post-treatment liver tissues were processed for histology activity index scoring and COX-2 immunohistochemical staining.

Immunohistochemistry

Liver tissues were fixed in 10% buffered formalin and embedded in paraffin. Sections, 5 μm thick, were cut and mounted on 3-aminopropyltriethoxysilane-coated slides (Marienfeld, Badmergentheim, Germany). Sections were de-paraffinized and re-hydrated in Tris-buffered saline. Endogenous peroxidase activity was blocked with 0.3% H2O2 for 10 min. Non-specific binding was blocked with 5% rabbit serum (DAKO, Glostrup, Denmark), and then incubated with antibody to COX-2 (1 : 100; Santa Cruz Biotechnology, Santa Cruz, CA, USA) in Tris-buffered saline containing 2% rabbit serum and 1% bovine serum albumin for 2 h. Tissues were incubated with the same buffer without the antibody as negative control. This was followed by biotinylated rabbit antigoat immunoglobulins (1 : 400; DAKO) and streptavidin/horseradish peroxidase complex (1 : 400; DAKO) incubation, both for 45 min. The colour was developed in diaminobenzidine solution (Sigma, St. Louis, MO, USA). The sections were then counter-stained with Mayer’s haematoxylin.

All stained specimens were scored by two independent investigators who were blind to the patient subgroups, treatments and histology activity index scores.5, 29 The scores were assigned on the basis of: (i) maximum intensity (i.e. the maximum level among all positive cells); (ii) dominant intensity (i.e. the level observed in the majority of positive cells); and (iii) extensiveness (by percentage population) of positively stained cells. Both maximum intensity and dominant intensity were scored on a scale of 0–3 (0, negative staining; 1, weak positive staining; 2, moderate positive staining; 3, strong positive staining), whereas extensiveness was scored on a scale of 0–4 (0, negative; 1, positive staining in 1–25% of cells; 2, positive staining in 26–50% of cells; 3, positive staining in 51–75% of cells; 4, positive staining in 76–100% of cells). The score of each specimen was the sum of the three parameters.

In situ hybridization

A 305-base pair complementary DNA fragment of human COX-2 cloned into pMOSBlue vector (Amersham, Buckinghamshire, UK) was used as a template to generate digoxigenin-labelled anti-sense or sense ribo-probe, as described previously.10 Anti-sense human glyceraldehyde-3-phosphate dehydrogenase ribo-probe was used as positive control. Paraffin-embedded tissue was cut into 5 μm sections and mounted on 3-aminopropyltriethoxysilane-coated slides. After de-paraffinization and re-hydration, the slides were soaked in 0.2 N hydrochloric acid for 20 min. This was followed by proteinase K (Sigma) digestion (10 μg/mL) in Tris–ethylenediaminetetra-acetic acid buffer at 37 °C for 9 min. The sections were further digested by 0.5 unit of RNase-free DNase in DNase buffer (Promega, Madison, WI, USA) at 37 °C for 30 min. Post-fixation was carried out in 4% formaldehyde (Sigma) in phosphate-buffered saline, pH 7.4, at 4 °C for 10 min. The sections were then hybridized with 1 ng/μL digoxigenin-labelled anti-sense or sense COX-2 ribo-probe in hybridization buffer containing 40% deionized formamide (BioRad, Hercules, CA, USA), 1 × Denhardt’s solution (Sigma), 10% dextran sulphate (Sigma), 2 × standard sodium citrate (Gibco-BRL, Gaithersburg, MD, USA) and 1 mg/mL yeast transfer RNA (Gibco-BRL), and incubated at 50 °C overnight in a humid chamber. After hybridization, the sections were washed sequentially by 2 × standard sodium citrate (twice) and 1 × standard sodium citrate (twice) at room temperature for 15 min each. Immunological detection was conducted using the DIG Nuclear Acid Detection Kit (Roche, Indianapolis, IN, USA). After colour development, the sections were counter-stained with nuclear fast red (Sigma).

Western blot analysis

Protein was extracted from frozen tissues as described previously.30 Tissues were homogenized in Tris–HCl (pH 7.4) buffer containing 0.5% Triton X-100 and protease inhibitor cocktail (Roche). Protein concentrations were measured by the method of Bradford (Bio-Rad). Fifty micrograms of protein was loaded per lane, separated by 10% sodium dodecyl sulphate polyacrylamide gel electrophoresis under reducing conditions, and transferred onto equilibrated polyvinylidene difluoride membrane (Amersham) by electro-blotting. Membranes were blocked by 5% non-fat dry milk. COX-2 protein was detected with a monoclonal antibody to COX-2 (Cayman, Ann Arbor, MI, USA) at a dilution of 1 : 1000 at 4 °C overnight. After secondary antibody incubation, enhanced chemiluminescence (Pierce, Rockford, IL, USA) was determined by exposure to X-ray film (Fuji, Dusseldorf, Germany). The equality of protein amount in all lanes was verified using reversible staining with Ponceau S.31 A cell lysate of human gastric adenocarcinoma cell line MKN 45, which highly expresses COX-2,32 was used as the positive control.

Reverse transcription polymerase chain reaction

Total RNA was isolated from frozen liver tissues using Trizol reagent (Gibco-BRL). RNA extracted from human gastric adenocarcinoma cell line MKN 45 was used as the positive control. Four micrograms of RNA from each sample was reverse transcribed using 1.5 U of avian myeloblastosis virus reverse transcriptase (Gibco-BRL) in a total reaction volume of 20 μL. One microlitre of reverse transcribed product was amplified by polymerase chain reaction using 0.75 U of Taq DNA polymerase (Gibco-BRL) and 10 pmol COX-2 forward and reverse primers. Each polymerase chain reaction cycle consisted of a denaturation step (96 °C, 24 s), an annealing step (58 °C, 48 s) and an elongation step (72 °C, 1 min). There were a total of 30 cycles, followed by an additional extension step (72 °C, 5 min). Ten picomoles of β-actin forward and reverse primers were added in the same multiplex polymerase chain reaction after seven cycles (23-cycle amplification) as an internal control for the efficiency of reverse transcription and the amount of RNA. The primer sequences (F, forward; R, reverse) have been described previously:33 COX-2, 5′-TTCAAATGAGATTGTGGGAAAATTGCT-3′ (F) and 5′-AGATCATCTCTGCCTGAGTATCTT-3′ (R); β-actin, 5′-CTAGAAGCATTTGCGGTGGACGATGGAGG-3′ (F) and 5′-TGACG GGGTCACCC ACACTGTGCCCATCTA-3′ (R). The sizes of the polymerase chain reaction products of COX-2 and β-actin were 305 and 654 base pairs, respectively.

The polymerase chain reaction products were separated on 1.5% agarose gel and saved as digital images (Uvigrab, UVItec, Cambridge, UK). The results were analysed using the UV Gel Documentation System (Uvigel, UVItec). The relative expression of COX-2 was determined by comparing the band intensity of COX-2 with that of β-actin for semiquantification.

Statistical analysis

A two-tailed Mann–Whitney U-test was used to compare results from the COX-2 immunohistochemical analysis and the necro-inflammatory and fibrosis scores of the different patient groups. Spearman’s correlation analysis was used to define the relationships between COX-2 expression and serum alanine aminotransferase and HBV DNA levels in chronic hepatitis B. Statistical significance was taken at P < 0.05. Bonferroni’s correction was performed for data with multiple comparisons.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Increased COX-2 expression in hepatocytes of chronic hepatitis B patients

Twenty-six HBeAg-positive and nine HBeAg-negative chronic hepatitis B patients were compared with 14 control subjects. The necro-inflammatory scores of HBeAg-positive chronic hepatitis B patients were significantly higher than those of HBeAg-negative chronic hepatitis B patients and controls (P < 0.005, Table 1). The fibrosis scores were not significantly different (P=1, Table 1). Both HBeAg-positive and HBeAg-negative chronic hepatitis B patients had significantly higher COX-2 expression than controls (P < 0.005, Table 2). There was no significant difference in COX-2 expression between HBeAg-positive and HBeAg-negative chronic hepatitis B patients (P=0.55, Table 2). Subgroup scores for COX-2 expression are summarized in Table 2.

Table 2.   Scores representing immunoreactivity for cyclooxygenase-2 (COX-2) Thumbnail image of

COX-2 expression in liver tissue of chronic hepatitis B patients was confined to hepatocytes diffusely distributed throughout the hepatic lobules (Figure 1A–C). Bile duct epithelium, vascular endothelium of the hepatic arteries, portal vein and sinusoids, Kupffer cells and inflammatory cells did not show significant COX-2 expression. Strong COX-2 expression was detected in chronic hepatitis B patients irrespective of HBeAg status and histology activity index scores (Figure 1A, B). Hepatocytes of non-alcoholic steato-hepatitis patients also expressed weak COX-2 protein in hepatic lobules (Figure 1D). In normal livers, very weak COX-2 expression was detected in isolated hepatocytes around central veins (Figure 1E). In situ hybridization using a specific anti-sense ribo-probe showed marked COX-2 messenger RNA expression in hepatocytes throughout the lobules of chronic hepatitis B patients (Figure 1F), with a pattern closely resembling that obtained in immunohistochemical studies. In sections hybridized with sense COX-2 ribo-probe, no positive signals were observed (Figure 1G).

image

Figure 1.  Immunohistochemical detection (A–E) and in situ hybridization (F,G) of cyclooxygenase-2 (COX-2) protein and messenger RNA, respectively, in liver biopsy specimens. In hepatitis B e antigen (HBeAg)-positive chronic hepatitis B patients (necro-inflammatory score=13), immunohistochemical staining of COX-2 revealed diffuse positivity in the hepatocytes throughout the lobules, while the inflammatory cells in the portal region were negative (A: original magnification, × 200). In HBeAg-negative chronic hepatitis B patients (necro-inflammatory score=3), similar diffuse staining for COX-2 in hepatocytes was observed (B: original magnification, × 200). A high power view demonstrated those hepatocytes showing variable staining intensity for COX-2. The Kupffer cells in the sinusoids were negative (C: original magnification, × 400). Hepatocytes of non-alcoholic steato-hepatitis patients expressed weak COX-2 protein in hepatic lobules (D: original magnification, × 200). In normal livers, weak COX-2 expression was detected in some hepatocytes around central veins (E: original magnification, × 200). In situ hybridization showed diffuse expression of COX-2 messenger RNA in hepatocytes throughout the lobules of chronic hepatitis B patients. Note that the expression pattern is consistent with that of immunohistochemical staining (F: original magnification, × 200). In the section hybridized with sense COX-2 ribo-probe, no positive signals were observed (G: original magnification, × 200).

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Strong expression of COX-2 enzyme in chronic hepatitis B patients was further confirmed by Western blot and reverse transcription polymerase chain reaction analysis. COX-2 protein could be demonstrated clearly in HBV-infected livers, but only weakly in control livers (Figure 2). In accordance with the Western analysis, reverse transcription polymerase chain reaction revealed strong COX-2 messenger RNA levels in HBV-infected livers. COX-2 messenger RNA could be barely detected in liver tissues from control subjects (Figure 3).

image

Figure 2.  Western blot analysis for cyclooxygenase-2 (COX-2) protein in tissue lysates of hepatitis B virus (HBV)-infected and normal livers. COX-2 protein expression was enhanced in HBV-infected livers. Lane G, human gastric adenocarcinoma cell line (MKN 45) as COX-2 positive control; lane H, HBV-infected livers; lane N, non-infected liver controls.

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image

Figure 3.  Reverse transcription polymerase chain reaction analysis for cyclooxygenase-2 (COX-2) messenger RNA expression in hepatitis B virus (HBV)-infected and normal livers. Strong COX-2 messenger RNA levels were demonstrated in HBV-infected livers. Lane M, 100-base pair DNA marker; lane G, human gastric adenocarcinoma cell line (MKN 45) as COX-2 positive control; lane H, HBV-infected livers; lane N, non-infected liver controls.

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The COX-2 immunoreactivity score did not correlate with serum alanine aminotransferase (R2=0.04, P=0.26; Figure 4A and Table 1) or HBV DNA (R2=0.10, P=0.11; Figure 4B and Table 1) levels.

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Figure 4.  Scatter plots illustrating the distribution of cyclooxygenase-2 (COX-2) immunoreactivity score with serum alanine aminotransferase (ALT) (A) and hepatitis B virus (HBV) DNA (B) levels in chronic hepatitis B patients. No correlation between COX-2 immunoreactivity score and serum ALT (R2=0.04, P=0.26) or HBV DNA (R2=0.10, P=0.11) levels was shown.

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Up-regulation of COX-2 persisted despite response to anti-viral therapy

Twelve patients received interferon or lamivudine therapy and responded favourably with a sustained HBeAg seroconversion (loss of HBeAg, emergence of anti-HBe antibody), disappearance of HBV DNA in serum and normalization of liver enzymes. There were significant reductions in necro-inflammatory activities, but not in fibrosis activities, in the paired biopsies after anti-viral treatment (Table 3). However, there was no appreciable change in COX-2 expression in both interferon-treated (P=0.55) and lamivudine-treated (P=0.62) chronic hepatitis B patients (Table 3 and Figure 5).

Table 3.   Effects of anti-viral therapy on cyclooxygenase-2 (COX-2) expression and histology activity index (HAI) Thumbnail image of
image

Figure 5.  Immunohistochemical detection of cyclooxygenase-2 (COX-2) protein in liver biopsy specimens from hepatitis B e antigen (HBeAg)-positive chronic hepatitis B patients pre- and post-treatment with interferon (A, pre-treatment; B, post-treatment; original magnification, × 200) or lamivudine (C, pre-treatment; D, post-treatment; original magnification, × 200). Although the inflammation subsided after treatment, both the pre- and post-treatment biopsies demonstrated diffuse immunostaining of COX-2 in the hepatocytes and no noticeable change in COX-2 expression could be observed.

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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

HBV and HCV are the most common causes of hepatocellular carcinoma around the world. Previous studies using immunohistochemistry have demonstrated increasing levels of COX-2 in patients with chronic hepatitis C, cirrhosis and hepatocellular carcinoma.5, 6 Prominent COX-2 expression has been detected in moderately and well-differentiated, but not poorly differentiated, tumour cells, suggesting that COX-2 may play a role in early hepatic carcinogenesis.5–7 In this study, we have demonstrated the up-regulation of COX-2 protein and messenger RNA levels in patients with chronic HBV infection compared with controls. Immunohistochemical analysis shows that COX-2 expression is confined to hepatocytes, as confirmed by in situ hybridization. This result is consistent with previous reports showing elevated COX-2 levels in patients with chronic hepatitis C.5, 6 These observations suggest that COX-2 may be involved in the pathogenesis of both HBV- and HCV-associated chronic liver diseases.

Does up-regulated COX-2 expression merely reflect an inflammatory process in the liver or a step towards cancer development? Results from the present study show that COX-2 expression, albeit induced by HBV-stimulated inflammation, persists after inflammation has subsided. Both HBeAg-positive and HBeAg-negative chronic hepatitis B patients showed higher levels of COX-2 expression compared to controls. More importantly, both HBeAg-positive and HBeAg-negative patients, despite showing significantly different inflammatory activities (reflected by necro-inflammatory scores), demonstrated a similar level of up-regulated COX-2 expression (Tables 1 and 2). Moreover, despite the marked decrease in inflammatory activity after successful anti-viral therapy, COX-2 expression in chronic hepatitis B patients showed no significant changes (Table 3). COX-2 expression was not correlated with serum alanine aminotransferase levels, a surrogate marker of necro-inflammatory activity (Figure 4A). These results strongly suggest that COX-2 expression in chronic hepatitis B patients does not merely reflect the degree of hepatic inflammation. This observation is in agreement with that of Agoff et al. who showed that COX-2 over-expression in ulcerative colitis-associated neoplasia cannot be simply explained by inflammatory activity alone.2 We have also demonstrated previously that, despite the eradication of Helicobacter pylori, which led to the resolution of mucosal inflammation, there was only a modest reduction of COX-2 in the gastric epithelium with intestinal metaplasia.10

The mechanism of COX-2 up-regulation in chronic hepatitis B patients is not entirely clear, but we have postulated three possible pathways based on existing evidence. Viral load and active viral replication are unlikely to be the main driving force, as COX-2 expression is not correlated with HBV DNA level and HBeAg status. It is well known that the viral genome integrates into the genome of patients in chronic HBV infection,34 and functional HBV x antigen (HBx) has been produced from such integrated genomes.35, 36 HBx can activate the Ras, Raf, mitogen-activated protein (MAP) kinase signalling cascade37 and the transcription factor nuclear factor (NF)-κB.38 A recent study has reported that HBx can also up-regulate the stress-activated protein kinase/c-jun-NH2-terminal kinase (SAPK/JNK) pathways.39 Because the activation of MAP kinase and the SAPK/JNK pathways can induce COX-2 expression,40–42 we postulate that HBx can up-regulate COX-2 in hepatocytes in chronic hepatitis B patients through these signalling pathways. In addition to its transcriptional transactivating properties, HBx can bind to and inactivate the tumour suppressor gene, p53.43 Because p53 can inhibit COX-2 transcription, inactivation of p53 after HBx binding may be involved in COX-2 over-expression in carcinogenesis.44 In agreement with this observation, a recent study has shown that p53 mutation is associated with COX-2 over-expression in gastric cancer.45 Thus, the HBx–p53 interaction may provide an additional pathway to induce COX-2 expression in chronic hepatitis B patients. A third possible pathway for COX-2 up-regulation in chronic hepatitis B patients involves the transforming growth factor-α/epidermal growth factor receptor system. In addition to COX-2, transforming growth factor-α expression is also elevated in hepatocytes in chronic viral hepatitis and well-differentiated hepatocellular carcinoma, but not in poorly differentiated hepatocellular carcinoma.29, 46 Similar to COX-2, transforming growth factor-α expression shows no significant difference in chronic hepatitis B patients before and after receiving interferon therapy.29 The fact that the activation of the transforming growth factor-α/epidermal growth factor receptor system in vitro induces COX-2 expression13, 47 further supports their relationship in chronic hepatitis B. Hepatocellular carcinoma has been shown to develop in transgenic mice over-expressing transforming growth factor-α.48 The interrelationship between COX-2 and the transforming growth factor-α/epidermal growth factor receptor system deserves further investigation.

In this study, we have demonstrated that COX-2 up-regulation in chronic hepatitis B patients persists despite HBeAg seroconversion, clearance of HBV DNA and normalization of liver enzymes after response to lamivudine or interferon therapy. It is therefore possible that the risk of development of hepatocellular carcinoma is not significantly reduced after successful anti-viral therapy. In fact, previous studies with long-term follow-up among interferon responders have yielded conflicting results with regard to the risk of hepatocellular carcinoma development.25–27 Data on the long-term effect of lamivudine in the prevention of hepatocellular carcinoma are lacking. A longer follow-up will be needed to substantiate the correlation between COX-2 expression and the development of hepatocellular carcinoma.

In conclusion, this study has demonstrated that COX-2 expression in hepatocytes is enhanced in chronic HBV infection. This elevation does not simply reflect inflammatory activity elicited by viral infection. Over-expression of COX-2 appears to persist even after successful anti-viral therapy. The HBV-mediated up-regulation of COX-2 may generate a long-term over-production of mutagenic and tumorigenic prostanoids in hepatocytes, leading to neoplasia. It would be interesting to study whether COX-2 inhibition by conventional non- steroidal anti-inflammatory drugs or COX-2-specific inhibitors is able to prevent hepatocellular carcinoma.

Acknowledgements

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

This study was supported by the Cheng Suen Man Shook Foundation for Hepatitis Studies.

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  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References
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