DDW 2007: Additional HCV Abstracts

 

Hepatobiliary Neoplasia: Clinical and Experimental

101. Serum glypican 3 after curative hepatectomy predicts early-phase recurrence of hepatocellular carcinoma. 

K. Watanabe; T. Asano; Y. Hippo; A. Watanabe; Y. Midorikawa; S. Ohnishi; T. Kodama; M. Omata; M. Makuuchi; H. Aburatani

 

Background:

Hepatocellular carcinoma (HCC) is an end-result of chronic infection with hepatitis B or C virus. High incidence of intrahepatic recurrence remains a major problem in the treatment of HCC. We have reported that serum glypican 3 is a potent biomarker especially for detection of well-to-moderately differentiated HCC. In the clinical course of HCC, curative treatment against tumor, preserved liver function and functional status of the patients are the important prognostic factor for patients with HCC. Meanwhile, the prognostic significance of glypican 3 has not been investigated in patients with HCC. Therefore, the aim of this study is to evaluate usefulness of glypican 3 as a biomarker of recurrent HCC after curative surgery.

 

Patients and Methods:

This study includes a total of 125 patients with HCC (64% HCV) consecutively diagnosed and treated with curative hepatectomy at University of Tokyo Hospital from October 2002 to December 2004. Curative hepatectomy was confirmed by negative surgical margin and absence of macroscopic vascular invasion. Serum glypican 3 was measured by sandwich ELISA. Patient characteristics, tumor stage, pathological grade, anatomic resection and serum tumor markers (alpha fetoprotein and glypican 3) were screened for their prognostic significance by univariate and multivariate analysis.

 

Results:

In 125 patients, 43 cases experienced recurrence in the follow up period (median; 15.5 months). Recurrence-free survival rate at 6 months and 1 year after surgery was 83.5% and 71.1%, respectively. In multivariate analysis, pathological diagnosis of microsatellite nodule (hazard ratio: HR 3.10, 95% confidence interval: CI 1.32-7.30, p < 0.01), microvascular infiltration (HR 2.29, 95%CI 1.15-4.53, p=0.01), prolongation of prothrombin time (HR 2.35, 95%CI 1.06-5.17, p=0.03) and serum glypican 3 measured postoperatively (glypican 3 > 0.3ng/ml: HR 2.46, 95%CI 1.04-5.82, p=0.03) were significant factors associated with recurrence. Patients with postoperative serum glypican 3 < 0.3 ng/ml of value was significantly higher recurrence-free rate than patients with glypican 3 0.3 ng/ml (91.9% vs 63.7% at 6 months and 80.2% vs 49.2% at one year, p < 0.001 by logrank test).

 

In conclusion, serum glypican 3 is useful as predictive marker of recurrence after curative surgical treatment of HCC.


Clinical Biliary Tract Disorders 

S1222. Incidence and Risk Factors for Hepatic and Extrahepatic Malignancies in Primary Biliary Cirrhosis in Japan. 

K. Sato; K. Hosonuma; M. Yanagisawa; T. Itikawa; N. Sohara; S. Kakizaki; H. Takagi; M. Mori

 

Background:

Hepatocellular carcinoma (HCC) have been increasingly reported as a complication of primary bliliary cirrhosis (PBC). However, no large case studies have focused on the detailed analysis of the incidence ratio and risk factors for hepatic and extrahepatic malignancies in PBC in Japan.

 

Methods:

Our aim was to analyze the incidence of malignancies in a large series of PBC patients in Japan; to compare the incidence with those obtained in the general population, as derived from the general cancer registry; and to clarify any possible adjunctive risk factors for malignancy. The comparison of the incidence of malignancies between the study group and the general population was obtained by the standardized incidence ratio (SIR), which is the ratio between the cases observed and the expected number of cases in the study group. A logistic regression analysis was performed to clarify the risk factors for hepatic or extrahepatic malignancies using laboratory data and questionnaires about hereditary predisposition and lifestyle habit.

 

Results:

The overall sample included 239 patients (29 males, 210 females). The mean age was 57 years (range 24-86); Liver biopsy was performed in 168 patients. One hundred and eight patients had Scheuer's stage 1, 33 had stage 2, 16 had stage 3, and 11 had stage 4. The follow-up period was average 89 months per person. Extrahepatic malignancies developed in 14 cases (5.9%), and HCC in eleven cases (5.2%) which were not always associated with cirrhosis. One case of HCC had positive HCV-RNA test but the non-cancerous liver tissue was consistent with PBC. The SIR for HCC was 6.0 (95%CI 3.0-10.7), whereas the SIR for top four extrahepatic malignancies, the colon, gastric, lung and breast cancer were 0.7 (95%CI 0.1-2), 0.6 (95%CI 0.1-1.7), 0.7 (95%CI 0.1-2.4), 1.2 (95%CI 0.1-4.3), respectively. Symptomatic PBC showing icterus, ascites, varices, or hepatic coma was an independent risk factor for HCC in overall PBC patients. When multivariate analysis was restricted to female patients, Age, HCV-RNA, symptomatic PBC or family history of HCC were independent risk factors for HCC. Diabetes mellitus, hyperlipidemia, symptomatic PBC, or preexistence of HCC were independent risk factors for extrahepatic malignancies in not only overall but also female patients.

 

Conclusions:

The incidence of HCC but not extrahepatic malignancies was significantly higher in PBC than those in the general population in Japan. We advocate that symptomatic PBC has to be regularly monitored for HCC and extrahepatic malignancies, and lifestyle-related diseases may play an important role on incidence of extrahepatic malignancies in PBC.

 


Clinical Hepatitis

S1225. Serum hepatocyte growth factor activator inhibitor type 1 is positively associated with hepatic fibrosis in patients with chronic hepatitis C virus infection. 

A. Moriuchi; H. Uto; S. Hasegawa; M. Oketani; Y. Takami; K. Kusumoto; K. Hayashi; A. Ido; T. Yamagishi; S. O. Stuver; H. Tsubouchi

 

Purpose:

Liver cirrhosis/fibrosis is characterized by excess production and deposition of extracellular matrix components, leading to tissue scarring and destruction of normal hepatic parenchyma. Hepatocyte growth factor (HGF) activator inhibitor type 1 (HAI-1) is a serine protease inhibitor that proteolytically cleaves and activates HGF. The mature membrane-form of HAI-1 can be proteolytically cleaved and released into the extracellular environment. Although several studies have investigated the anti-fibrotic effect of HGF, how the HGF/HAI-1 system is involved in hepatocyte repair and hepatic fibrosis has not been fully clarified. The aim of this study is to elucidate the clinical significance of serum HAI-1 in patients with chronic hepatitis C virus (HCV) infection.

 

Methods:

A total of 454 subjects with chronic HCV infection were included in this study. Chronic HCV infection was confirmed by the presence of HCV antibodies and HCV RNA. An additional 89 chronic hepatitis C (CHC) patients who underwent liver biopsy were also analyzed. Serum HAI-1 levels (ng/ml) were assessed by enzyme-linked immunosorbent assay (ELISA). In addition to HAI-1, serum levels of total HGF, activated HGF, platelet count, AST, HCV core protein and hepatic fibrosis marker, type IV collagen 7S (IVcol7s) and hyarulonic acid (HA) were also evaluated.

 

Results:

Mean HAI-1 serum levels were 7.5±2.3 in the healthy donors (n=52). In the 454 HCV-infected subjects, serum HAI-1 levels were positively correlated with IVcol7s (r=0.73, p<0.001) and HA (r=0.42, p<0.001). However, there was no correlation between HAI-1 levels and total HGF, activated HGF, AST, or HCV core protein levels. In the 89 patients with CHC, serum HAI-1 levels significantly increased with the severity of liver fibrosis [F1:12.5±7.6 (n=38); F2:18.2±9.6 (n=28); F3:26.4±11.0 (n=11); F4:30.1±13.0 (n=12)] (p<0.001). In addition, the nine patients who remained responsive to interferon therapy had significantly decreased mean HAI-1 levels (-7.6±2.0) over the seven-year follow-up period, whereas in the 11 untreated patients, mean HAI-1 levels increased during this observation period (18.0±3.9) (p<0.001).

 

Conclusions:

HAI-1 is associated with hepatic fibrosis but not with HGF levels. Serum HAI-1 levels may serve as a non-invasive, clinical marker for hepatic fibrosis in HCV-infected patients. HAI-1 levels could also serve as a surrogate marker for the antifibrotic effects of HCV treatment.

 


Esophagus

S1340. Clinical Predictors for Recurrence of Esophageal Varices after Obliteration by Endoscopic Band Ligation. 

V. Chandrasekhara; J. Yepuri; J. Sreenarasimhaiah

 

Background:

There is no universal standard of care for endoscopic follow-up after obliteration of esophageal varices by esophageal variceal ligation (EVL).

 

Aim:

Identify clinical predictive factors and rate of varices recurrence following EVL.

 

Methods:

All patients presenting to Parkland Memorial Hospital for EVL were evaluated since October 2005. Those with prior bleeding gastric varices, portal decompressive procedures, or variceal sclerotherapy were excluded. Patients consented to repeate EVL every 3 weeks until obliteration, and then repeat endoscopy every 3 months for 1 year. Clinical and laboratory parameters were monitored continuously over this period.

 

Results:  

Of 31 patients enrolled to date, 35% were Caucasian and 65% were Hispanic. 71% of patients were male and 29% female with mean age of 48 years. At the initial time of EVL, 39% were still consuming alcohol. The cause of cirrhosis was alcohol alone (26%), hepatitis C (HCV) alone (26%), combined HCV and alcohol (23%), cryptogenic (9%), nonalcoholic fatty liver (3.2%), primary biliary cirrhosis (3.2%), combined hepatitis B (HBV)/HCV (3.2%), and combined alcohol/HBV/HCV (3.2%). Noncompliance with the EVL protocol was noted in 10/31 (32%) patients. Of these, 4/10 (40%)were Caucasian, 6/10 (60%) were Hispanic, 3/10 (30%) were non-English speaking, and 8/10 (80%)were actively consuming alcohol. Variceal rebleeding was noted in 8/31 (26%) within 1 year of initial EVL. Of these, 4/8 (50%) were non-compliant with the EVL protocol. Alcoholic cirrhosis was noted in 5 of 8 (62.5%) patients. Moreover, 4/5 (80%) were still actively drinking. For all 31 patients, the mean MELD score was 13.3 and the mean CPT score was 8.1. The mean MELD score was 16.1 for those with variceal rebleeding and 12.3 for those without rebleeding (p=0.013). The mean INR in the rebleeding group was 1.64 and in the non-rebleeding group was 1.37 (p=0.017). The average age for the two groups was 49.2 and 47.5 years, respectively. There has been one death thus far due to a bleeding gastric lesion.

 

Conclusions:  

Recurrence of variceal bleeding after EVL obliteration occurred in those patients with a higher MELD score and specifically a higher INR level when compared to those who did not bleed. Patients with alcoholic cirrhosis had a higher rate of rebleeding. There was no correlation with patient age. Rebleeding was higher in those who were noncompliant with the EVL protocol including patients who were non-English speaking as well as those with active alcohol consumption. Therefore, efforts needed to be directed to facilitating patient compliance with medical care and abstinence from alcohol.


Clinical Hepatitis: Advances in the basis for and markers of progressive disease

585. Elevated levels of circulating mesenchymal stem cells in patients with chronic hepatitis B and C

J. A. Abrams; V. Jacob; J. S. Manavalan; T. C. Wang; S. Wang

 

Background:

Adult stem cells play a crucial role in the repair and maintenance of tissues. There are three major populations of adult stem cells: mesenchymal stem cells (MSC), endothelial progenitor cells (EPC), and hematopoietic stem cells (HSC). Mesenchymal stem cells generate fibrous connective tissue, bone, cartilage, and the reticular network that supports blood cell formation. The pathways that lead to hepatic fibrosis in chronic hepatitis are not completely understood, and the recruitment of MSCs and other progenitor cells may be involved in this process. We conducted an exploratory analysis of circulating progenitor cells in patients with chronic viral hepatitis.

 

Methods:

Peripheral blood mononuclear cells (PBMCs) were isolated from 7 patients with chronic hepatitis B or C without cirrhosis and from 7 healthy controls. Data were collected regarding age, gender, and stage of fibrosis on liver biopsy. Quantification of circulating progenitor cells was performed by flow cytometry. PBMCs were labeled with fluorochrome conjugated monoclonal antibodies against CD45, CD34, CD133, VEGFR2, CD117, and CD44, and incubated and washed prior to 6-color analysis using a LSR II flow cytometer. EPCs were defined as CD34+ CD133+ VEGFR2+, HSCs as CD45+ CD34+ CD117+, and MSCs as CD45- CD44+. Counts were expressed as number of progenitor cells per 10,000 mononuclear cells.

 

Results:

Patients with chronic hepatitis B or C had a significantly higher mean number of MSCs per 10,000 mononuclear cells than healthy controls (68.7 vs. 38.9, p=0.04) (Table 1). There was no difference between patients with chronic viral hepatitis and controls with respect to number of circulating EPCs (2.8 vs. 2.9, p=0.91) or HSCs (6.7 vs. 5.5, p=0.50). There was no significant correlation between number of circulating MSCs and age, gender, or stage of fibrosis on liver biopsy.

 

Conclusions:

Mesenchymal stem cells are present in significantly higher levels in serum from patients with chronic hepatitis B and C than from healthy controls. It is possible that recruitment of MSCs plays an important role in the development of hepatic fibrosis. Further studies should be aimed at defining the role of MSCs in hepatic fibrosis and identifying the signaling mechanisms that trigger their recruitment.

 

Table 1. Mean number of circulating progenitor cells per 10,000 mononuclear cells in patients with hepatitis B or C and in healthy controls. Standard deviations in parentheses.

 

 

Hepatitis B or C

Controls

p-value

MSC

68.7 (28.9)

38.9 (18.1)

0.04

EPC

2.8 (1.3)

2.9 (2.4)

0.91

HSC

6.7 (3.5)

5.5 (2.9)

0.50

 


Clinical Portal Hypertension Poster Session

T1039. Impact of prophylactic antibiotics in cirrhotic patients presenting with sepsis. 

L. Lim; S. Lim; D. Sutedja; Y. Dan; Y. Lee; C. Wai

 

Aim:

Cirrhotic patients are prone to infection. Prophylactic antibiotics after an episode of spontaneous bacterial peritonitis (SBP) could potentially select out resistant organisms, such as extended spectrum beta-lactamase (ESBL)-producing organisms. We aimed to investigate the characteristics and outcomes of patients with cirrhosis who were admitted for sepsis of any source in our institution, which is a tertiary university liver transplant center.

 

Methods:

All patients with liver cirrhosis admitted to our institution from March 2004 to August 2006 for sepsis, who had documented organisms in any body fluid cultures, including blood, urine, ascites, sputum or pleural fluid, were retrospectively studied.

 

Results:

47 patients with a total of 63 admissions (16 for SBP, 15 for urinary tract infection) were studied: 26 (55%) were male, aged 65.3±1.6 years, whose major etiology of cirrhosis were cryptogenic cirrhosis (32%), chronic hepatitis B (26%), and alcoholic liver disease (26%). 13 (28%) had hepatocellular carcinoma. Of the 63 admissions, fever and abdominal pain was the presenting complaint in 36 (57%) and 26 (41%) respectively, with mean white cell count at 9.1±0.9 x10(9)/L, and mean C-reactive protein at 6.8±0.7mg/dL. Organisms isolated in cultures were, in decreasing order of frequency: Escherichia coli (22%), Klebseilla pneumoniae (18%), ESBL-producing Escherichia coli (13%), ESBL-producing Klebseilla pneumoniae (8%). Patients on prophylactic ciprofloxacin for prior SBP were more likely to harbor ciprofloxacin-resistant organisms (90% vs. 43%, p=0.007), and ESBL-producing organisms (50% vs. 15%, p=0.012) than those without. 5 of 6 (83%) episodes due to pneumonia compared with 12 of 57 (21%) episodes with infection other than pneumonia resulted in death (p=0.001). 16 of 37 (43%) septic episodes involving Child’s C patients resulted in death, compared with 1 of 26 (4%) involving Child’s A or B patients (p=0.001). The mean Child’s score (12.9±0.4 vs 9.4±0.4, p= 0.000) and MELD score (24.4±1.9 vs 17.3±0.9, p=0.000) were higher in those who died than those who survived. Using multivariate analysis, high Child-Pugh score (OR=0.579, p=0.021) and pneumonia (OR=12.6, p=0.043) had significant correlations with mortality.

 

Conclusions:

Sepsis, in particular pneumonia, was an important cause of mortality in cirrhotics, especially in those with high Child-Pugh score. Cirrhotics who were on antibiotic prophylaxis were more likely to be infected with ESBL-producing organisms, and should be started empirically on second line antibiotics such as carbapenems upon diagnosis.


Steatohepatitis: Clinical

 

T1064. Health Utility Assessment Using SF-6D and Health Utility Index (HUI) in Patients with Chronic Viral Hepatitis and Non-alcoholic Fatty Liver Disaese (NAFLD). 

 A. Dan; J. Kallman; R. Srivastava; Z. Younoszai; A. Kim; Z. M. Younossi

 

Introduction:

Assessment of health-related quality of life (HRQL) and health utilities have become important aspects of clinical research. Patient-derived utility-adjustments are frequently used in economic analysis. Although SF-36 is widely used as a generic HRQL instrument for patients with liver disease, extensive data on the health utilities of patients with liver disease are not available. Recently, SF-6D has been developed to obtain utility scores from the SF-36 questionnaire.

 

Aim:

To assess health utilities of patients with chronic liver disease using two utilities assessments (SF-6D and HUI). Methods: Patients with chronic liver disease were identified from our HRQL databases with available clinical data, SF-36, and HUI scores. SF-6D scores were calculated from SF-36.

 

Results:

140 patients were included[(42% female, age 49.4±11.2, 36% Hepatitis B, 29% Hepatitis C, 24% cholestatic liver disease, and 11% NAFLD. Analyses indicate that Hepatitis B patients had the best HRQL as measured by the SF-6D subscales (only mental health scores did not vary by health condition), while patients with Hepatitis C and cholestatic liver disease had similar HRQL, and those with NAFLD/other had the poorest HRQL (Table 1). HUI scores did not differ as much as SF-6D scores, but those scales that varied also indicated that those with other liver diseases had the poorest quality of life. Female patients experienced poorer HRQL as measured by the pain, social functioning and vitality SF-6D subscales and six of the HUI subscales. Older age was also moderately associated with many of the HRQL subscales.

 

Conclusion:

SF-36 with SF-6D scoring provides not only a generic assessment of HRQL but also a utility score that can be used for economic analysis.

 

 

Cholestatic Liver Disease

Hepatitis C

Hepatitis B

NAFLD/Other

Possible Range

SF-6D

 

 

 

 

 

Physical Functioning

4.5 (1.2)

4.4 (1.6)

5.0 (1.2)

3.4 (1.9) **

1-6

Role limitation

1.6 (.5)

1.5 (.5)

1.3 (.4)

1.8 (.4) ***

1-2

Social Functioning

3.8 (1.2)

3.6 (1.3)

4.4 (.9)

3.4 (1.5) **

1-5

Bodily Pain

3.9 (1.3)

3.9 (1.2)

5.0 (1.3)

3.9 (1.2) ***

1-6

Mental Health

4.5 (1.0)

4.3 (1.1)

4.6 (.8)

4.1 (1.1)

1-5

Vitality

3.4 (1.5)

3.3 (1.5)

4.3 (1.0)

3.1 (1.3) ***

1-5

HUI- Mark 2

 

 

 

 

 

Overall

0.8 (.2)

0.7 (.2)

0.9 (.2)

0.7 (.2) **

0-1

Cognition

0.9 (.1)

0.9 (.1)

1.0 (.1)

0.9 (.1)

0-1

Emotion

0.9 (.1)

0.9 (.2)

0.9 (.1)

0.9 (.2)

0-1

Mobility

1.0 (.1)

0.9 (.1)

1.0 (.0)

0.9 (.2) *

0-1

Pain

0.9 (.2)

0.8 (.2)

0.9 (.2)

0.9 (.3)

0-1

Self Care

1.0 (.2)

1.0 (.2)

1.0 (.1)

0.9 (.3)

0-1

Sensation

0.8 (.1)

0.8 (.2)

0.9 (.1)

0.8 (.1)

0-1

 

*p<.05 **p<.01 ***p<.001 Note: Higher values on utility scale scores as presented above reflect higher HRQL.

 


T1084. Elevated Serum Alanine Aminotransferase (ALT) Activity Does Not Increase Cardiovascular Disease (CVD) Mortality in the United States Population. 

C. E. Ruhl; J. E. Everhart

 

Introduction:

A concern exists that nonalcoholic fatty liver disease may increase the risk of CVD independently of metabolic syndrome components. In a prospective, national, population-based sample, we examined whether elevated ALT was associated with increased risk of all-cause and CVD mortality.

 

Methods:

Data were analyzed from 14,950 adult participants in the third U.S. National Health and Nutrition Examination Survey, 1988-94, who were negative for markers of viral hepatitis B and C. Abnormal ALT activity was defined as >40 U/L in men or >31 U/L in women. Participants were followed through 2000 for all-cause and CVD mortality as identified by death certificate underlying-cause diagnoses. Hazard rate ratios (HR) for mortality outcomes were calculated using Cox proportional hazards analysis to adjust for CVD risk factors. All analyses incorporated sample weights and the design effects of the survey.

 

Results:

The prevalence of elevated ALT was 5.3%. During the up to 12 years of follow-up, the cumulative mortality was 13.9% (2,189 deaths) from all causes and 4.2% (665 deaths) from CVD. Participants with an elevated ALT had a non-statistically significant 40% higher risk of all-cause mortality both in age-adjusted analysis (HR=1.4, 95% confidence interval (CI)=0.90-2.1) and after adjusting for multiple factors (HR=1.4, 95% CI=0.88-2.3). Interestingly, the risk of CVD mortality was not increased among persons with an elevated ALT in either age-adjusted (HR=1.1, 95% CI=0.44-2.8) or multivariate-adjusted analysis (HR=1.0, 95% CI=0.37-2.9).

 

Conclusion:

In the U.S. population, elevated ALT may be associated with an overall increase in mortality, but does not predict an increased risk of CVD mortality.

 


Epidemiology

W1044. Digestive Diseases in the Native American Population of the U.S. Central Plains: Disparities in Prevalence, Screening, and Treatment. 

B. Kizer; H. Lazarte; G. Haynatzki; R. Patel; A. Dajani; J. O'Brien

 

Background:

Little is known about the prevalence of digestive diseases among the Native American population from the United States central plains. The Fred Leroy Clinic of Omaha, NE, provides free medical care exclusively for over 4,000 Native Americans from 70 tribes throughout the Dakotas, Nebraska, and Kansas.

 

Methods:

A retrospective study of 449 randomly selected Fred Leroy charts. Each patient chart was reviewed for medical and social history, and the documentation of the prevalence, clinical presentation, and treatments of the ten most common digestive diseases in the United States. Data was also collected on health screening for these conditions. The one-sample z-test for proportions was used in all statistical analyses.

 

Results:

449 patient charts were reviewed. 138 patients were male; 311 were female. 330 patients were 19 to 50 years old; 119 patients were aged 50 to 79.

 

The Body Mass Index could be calculated for all but 2 patients. 113 patients were overweight (BMI 25 to < 30), 131 were obese (BMI 30 to < 35), and 137 were morbidly obese (BMI > 35). The prevalence in this study population of overweight / obesity is 85.3 % (p<0.00001, U.S. prevalence of 65%), the prevalence of obesity and morbid obesity is 60.0% (p<0.00001, U.S. prevalence of 30%).

 

No statistical difference existed between the study population and the U.S. population with regards to the prevalence of colorectal cancer (CRC). However, of 123 persons eligible for CRC screening, only 14 (11.4%, p<0.00001) were offered screening - well below the 42.5% of eligible U.S. patients who underwent screening in year 2000.

 

61 patients (13.6%) have a history of cholelithiasis. No statistically significant difference exists between the prevalence of cholelithiasis in this Native American population and the U.S. prevalence of 10-15% (p<0.00001).

 

27 patients (6%, p<0.00001) have a history of Hepatitis C infection (U.S. prevalence 1.7%).

 

Conclusion:

This is the first study looking at the prevalence of digestive diseases in the Native American population of the U.S. central plains. This population has a dramatically higher prevalence of overweight/obesity and hepatitis C compared to the general U.S. population. This population also lacks access to age and family history-appropriate colorectal cancer screening, which is a standard-of-care in the United States. The prevalence of cholelithiasis in this population is less than the rate reported in the Pima Indians, suggesting geographic variability. This indicates that data on prevalence of digestive diseases from Native Americans of the southwest United States cannot be applied to all Native American populations.

 


Immune Response

280. The Envelope Protein of the Hepatitis C virus (E2) inhibits IL-2 Secretion from T lymphocytes. 

D. Petrovic; L. Golden-Mason; S. Mitchell; Y. Volkov; C. O'Farrelly; D. Kelleher; A. Long

 

Introduction:

T-cell activation is one of the primary responses of the adaptive immune system to pathogens, such as Hepatitis C virus (HCV). Recognition of antigen, via the T cell receptor, initiates signal transduction and the resultant production of IL-2 is central to this response. We have previously demonstrated that the PKC-B enzyme is critical for the process of IL-2 secretion but not mRNA production (Long; et al). More recently we have shown that the Envelope Protein (E2) of the HCV, through its interaction with CD81, modulates PKC-B activity by stimulating its sequestration in lipid rafts, preventing T cell migration (Volkov; et al).

 

Aim:

The aim of this study was to investigate the effect of HCV E2 on T cell IL-2 production.

 

Results:

Analysis of cytokine levels in extracts of snap-frozen liver biopsies from individuals infected with HCV, measured by ELISA, revealed significantly lower levels of IL-2 in these patients when compared to those with other inflammatory liver diseases, such as ALD and PBC (IL-2 levels in ng/100mg protein: 5 ± 0.93 (HCV) vs 25 ± 2.9 (ALD), p<0.027 vs 20 ± 2 (PBC), p<0.003). In a HuT 78 T cell line model, pretreatment of cells with E2 substantially attenuated PMA stimulated IL-2 production (IL-2 levels in pg/ml: PMA stimulated cells 889.3 ± 19.1 vs E2/PMA stimulation 105.9 ± 0.2, p<0.016, (resting cells 20 ± 0.9)). This was also observed in PBMCs, obtained from healthy donors, stimulated with either PMA/Ionomycin or anti CD3 / anti CD28 antibodies (IL-2 levels in pg/ml: antiCD3/antiCD28 1825 ± 58.3 vs E2/antiCD3/antiCD28 53 ± 1.5, p<0.022, (resting cells 27.4 ± 1.4)). In contrast, cells pretreated with viral core protein C22 and NS3 protein C33 did not affect IL-2 secretion, confirming specificity of E2 in inhibition of IL-2 secretion. Inhibition of IL-2 was at the level of secretion and not transcription, as demonstrated by Real Time PCR (IL-2 mRNA level in PMA stimulated cells increased 2.07 fold over resting cells, p<0.01 while it increased IL-2 mRNA 2.19 fold in the presence of E2, p<0.0005). In addition, intracellular IL-2 was detectable by immunofluorescence in PMA activated cells pretreated with E2. Treatment of the HuT 78 cells with the lipid raft disruptor methyl-β-cyclodextrin (MCD) reversed the ability of E2 to inhibit IL-2 secretion.

 

Introduction:

These data suggest that targeting of PKC-B to the lipid raft microdomains in the T cell membrane, following E2 - CD81 interaction, disrupts the association of PKC-B with the microtubule cytoskeleton, thereby inhibiting export of IL-2. E2-mediated inhibition of IL-2 secretion represents a mechanism whereby HCV can undermine the human immune response to establish persistent infection and chronic disease.