POSTERSare associated with antiviral effector T-cell dysfunction, which influences on T-cell exhaustion and persistent viral infection.These PD-1 and CTLA-4 are up-regulated in chronic viral infection such as chronic hepatitis C, chronic hepatitis B and human immunodeficiency virus infection but there is no report about the role of PD-1 and CTLA-4 in patients with symptomatic acute hepatitis A. We investigated the expression of PD-1 and CTLA-4 during symptomatic and convalescent phases of acute hepatitis A. Methods: Seven patients with symptomatic acute hepatitis A, 5 patients with non-viral acute toxic hepatitis were enrolled for detection of PD-1 and CTLA-4 on T-cell subset of peripheral blood mononuclear cells (PBMC) isolated from these subjects during symptomatic and convalescent phases by flow cytometry.Five as healthy control were also examined for comparison with these patients.Results: Symptomatic acute hepatitis A showed significant increase of PD-1 and CTLA-4 expression compared to non-viral acute toxic hepatitis or healthy control (PD-1; 18.3±15.7%vs. 3.7±3.0%vs. 1.6±1.8%,p < 0.05, CTLA-4; 23.5±12.0%vs. 6.1±1.2% vs. 5.9±1.2%,p < 0.05) (median ± SD).In addition, highly expressed PD-1 and CTLA-4 were dramatically decreased in convalescent phase of acute hepatitis A. Conclusions: In acute hepatitis A, PD-1 and CTLA-4 are upregulated during symptomatic phase then down-regulated after recovery.This changing pattern of PD-1 and CTLA-4 expression was not seen in non-viral acute toxic hepatitis.Our findings suggest that PD-1 and CTLA-4 have protective effect as inhibitory molecules to suppress cytotoxic T-cells which induce destruction of viral infected hepatocytes.

Introduction:Progressive hepatic fibrosis is a common feature of chronic hepatitis C. Natural Killer (NK) cells represent a major component of intra-hepatic lymphocytes and have been shown to be important for the early control and natural course of HCV infection.Moreover, in mouse models it has been shown that natural killer cells can attenuate liver fibrosis via killing of activated hepatic stellate cells (HSC) in a NKG2D and TRAIL dependent manner.Here, we analyzed the interactions of human NK cells from HCVpositive patients with activated primary HSC and the potential effects of interferon-a (IFN-a) treatment.Material and methods: NK cells from untreated HCV-RNA(+) patients (n = 11), interferon-a (IFN-a) treated patients (n = 10) and healthy controls (n = 12) were co-incubated with activated primary HSC (ScienCell).NK cells from healthy persons were incubated in the presence or absence of IFN-a (25 IU/ml).Cytotoxic activity of NK cells was studied using the CD107a assay.In addition, INF-g and TNFa production of NK cells was measured by FACS analysis.Induction of HSC apoptosis (active Caspase-3) was analyzed flowcytometrically.Results: Analyzing cytolytic activity of NK cells following coincubation with HSC only discrete CD107a expression could be observed on both NK cells from HCV(+) patients and healthy controls ([mean±SEM: 5.7±0.7% vs. 4.7±0.8%;p = n.s.).Furthermore, only negligible secretion of IFN-g and TNF-a could be observed.However, NK cells from untreated HCV-infected patients were significantly more effective in induction of HSC apoptosis (20.8±2.1%)than NK cells from healthy controls (6.2±0.6%).Of note, NK cells from IFN-a/RBV treated HCV(+) patients displayed an even stronger capability to kill HSC (26.9±4.3%).In vitro stimulation of NK cells with IFN-a resulted in a significant up-regulation of TRAIL and was associated with increased killing of HSC as compared to un-stimulated NK cells.This effect could be blocked with both NKG2D-and TRAIL-specific antibodies.Conclusion: NK cells from HCV-infected patients are highly efficient in inducing apoptosis of activated hepatic stellate cells.This function of NK cell is increased following IFN-a treatment.Thus, NK cells may play an important anti-fibrotic role in chronic hepatitis C.

Background:The factors affecting response to HCV-specific therapy are only partly understood at the moment.However, three recent reports demonstrated a genetic polymorphism near the IL28B gene, encoding interferon-l-3, to be strongly associated with response to treatment with pegylated interferon-a and ribavirin in HCV mono-infected patients.Since co-infection with the hepatitis C virus (HCV) constitutes a major health problem worldwide in HIVpositive patients, we studied whether the IL28B gene polymorphism (rs12979860) also affects treatment outcomes in HCV/HIV coinfected patients.Methods: IL28B genotypes were determined in 192 HCV/HIV coinfected patients treated with pegylated interferon-a, including 74 patients with acute and 118 with chronic hepatitis C. Rates of sustained virological responses (SVR) were compared in patients carrying different genotypes.As a control, 137 healthy and 199 HCV mono-infected subjects were studied.Results: IL28B genotype distribution did not differ significantly between HIV-positive patients with acute versus chronic HCV coinfection and the two other study groups.In HIV/HCV co-infection carriers of C/C genotype had significantly lower HIV loads than patients with other genotypes (41 ×10 3 copies/ml [0.11-341]) vs. 95.9×10 3 copies/ml [0.12-1371], p = 0.038 C/C versus C/T and T/T).However, this difference must be attributed exclusively to HIV-positive patients with chronic hepatitis C (p = 0.011).Unlike HIV serum levels, HCV loads were significantly higher in carriers of the C/C genotype (12.1×10 6 IU/ml [<0.1-124] vs. 4.8×10 6 IU/ml [<0.1-46]; p = 0.0026).Of note, HCV/HIV patients with the homozygous C/C genotype had significantly higher SVR rates than patients with other genotypes (C/T and T/T combined) (58.2% vs. 40.6%;OR 1.6; p = 0.04).However, this effect reached statistical significance only in the HIV-positive subgroup of patients with chronic hepatitis C (50% versus 29%; P = 0.02; OR 2.4) but not in the subgroup with acute hepatitis C (71.4% versus 59%; n.s.). Conclusion:The IL28B genetic polymorphism has only limited effect on treatment-induced clearance of hepatitis C virus in HCV/HIV co-infected patients.This is in strong contrast to observations in HCV mono-infected patients and may suggest interference of HIV with the interferon-l signaling pathways.

The most notable changes occurred mainly in interferon-stimulated genes.Treatment response could be predicted with a molecular gene signature.The genes included in the signature encode molecules secreted in the serum and provide a logical functional approach for the development of serum markers to predict response to treatment.

Introduction: Dys-regulated function of natural killer (NK) cells has been proposed as a mechanism contributing to viral persistence in hepatitis C virus (HCV) infection and HCV-associated liver damage.Recent data indicate that members of the signaling lymphocytic activating molecule (SLAM) family receptors (i.e.NTB-A, CRACC, and 2B4) and their respective ligands NTB-A, CRACC, and CD48 have important functions in NK cell biology.However, in hepatitis C expression of these molecules has not been studied in great detail.Methods: NK cells from 54 patients with chronic hepatitis C, 26 subjects who had achieved a sustained virological response after after antiviral therapy (n = 26), and 47 healthy individuals were analysed by flow cytometry.Results: Expression of 2B4 and NTBA was not altered in HCV infection.However, expression of CRACC was significantly downregulated on CD56 bright NK cells in chronic hepatitis C (mean fluorescence index; HCV vs. healthy controls: 7.3 vs. 8.8, p < 0.01).In contrast to CRACC, expression of the ligand CD48 was significantly higher in the NK cell subsets from HCV-positive patients than in healthy controls (CD56 dim : 32.92 vs. 26.37,p = 0.02; CD56 bright : 12.79 vs. 9.6, p < 0.01).Interestingly, expression of CD48 on CD56 dim NK cells correlated positively with elevated aminotransferase levels (ALT: p = 0.02; AST: p < 0.01).Successful anti-viral treatment was associated with normalization of CD48 levels.Conclusions: This study demonstrates altered expression of the SLAM receptor CRACC and CD48 on NK cells in chronic hepatitis C.This phenotype may contribute to dys-regulated function of NK cells in HCV infections.

Background: NIM 811 is a cyclophilin inhibitor with substantially lower immunosuppressive activity compared with cyclosporine A. NIM811 shows potent in vitro anti-HCV activity and is currently being evaluated in a two-week proof-of-concept study for antiviral activity in HCV genotype-1 patients.Methods: Single oral doses of NIM811 (50-1600 mg) were administered to 40 healthy volunteers.Subjects were enrolled in 5 sequential dose groups (50, 150, 400, 800 or 1600 mg); 6 active and 2 placebo subjects per group, and one food effect group (150 mg).Multiple oral doses of NIM811 (25-600 mg QD or bid) were administered to 72 HCV genotype-1 patients for 14 days.Patients were enrolled in 6 sequential dose groups (25, 75, 100, 200, 400 or 600 mg); 9 active and 3 placebo patients per group.PK and safety assessments were conducted throughout the study period.Results: There were no SAEs, and no drug-related AEs in either lab results or clinical evaluation with single dose study in the healthy subjects.The compound was also well tolerated at all doses with multiple doses in the HCV-infected patients.Mild, clinically non-significant elevations of bilirubin and declines in platelet numbers were observed in the 400 and 600 mg bid groups.Blood exposure to NIM811 increased with increasing doses of NIM811.Terminal half-life of NIM811was around 12-20 hrs.Following multiple doses, NIM811 accumulated slightly upon reaching steady state (accumulation factor ~1.2).The inter-subject variability of NIM811 blood levels was relatively low (<30%).Food delayed NIM811 absorption (~2 hrs) and decreased Cmax and AUC (56% and 20%, respectively).Conclusions: NIM811 was well tolerated in healthy subjects and HCVinfected patients following single and multiple oral administration up to 1600 mg.The PK profile indicated good exposure to NIM811.Based on these results, twice a day dose regimen is expected to provide more efficient C trough concentrations for clinical antiviral efficacy.

Introduction:HLA-E is a non-classical MHC class I molecule with two allelic variants (HLA-E R and HLA-E G ). Recently, we demonstrated chronic hepatitis C to be associated with increased HLA-E expression and identified peptide HCV core aa35-44 as a ligand for HLA-E that stabilizes HLA-E surface expression, resulting in impaired NK cell function.Beyond interactions with NK cells, HLA-E has been suggested to also represent a ligand recognized by the TCR expressed by CD8(+) T cells.Here, we describe HLA-E restricted recognition of peptide HCV core aa35-44 by human CD8(+) T cells and its potential functional relevance in hepatitis C. Methods: HLA-E restricted T cell responses to HCV core aa35-44 were studied in 46 HCV infected patients and 11 healthy controls using HLA-E-transfected K562 cells for antigen presentation and the IFN-gamma ELISpot to detect responses.The HLA-E allelic polymorphism was studied in 201 patients with chronic hepatitis C and 140 healthy controls using lightcycler real time PCR.Results: HLA-E restricted HCV-specific CD8+ T-cell responses could be detected in 12/46 HCV patients (26%) but in none of the controls.Importantly, patients with HLA-E-restricted IFN-g responses displayed significantly lower HCV loads than patients without such a response (4.3×10 6 [3.5×10 5 -16.5×10 6 ] vs. 1.9×10 6 [1.1×10 5 -8.6×10 6 ]; p = 0.034).Moreover, HLA-E restricted T cell responses were associated with significantly increased serum levels of ; p = 0.007) and ; p = 0.012).Interestingly, the frequency of HLA-E restricted responses was significantly higher in patients homozygous for the HLA-E R allele as compared to carriers of the other allele (7/14 [50%] vs. 5/32 [15.6%]; p = 0.027).Moreover, patients with a chronic hepatitis C were significantly less likely to carry a homozygous HLA-E R genotype (42/201 [20.9%]) as compared to healthy subjects (43/140 [30.7%]; p = 0.042). Conclusion:Here, we show that CD8+ T cells can recognize a HCVderived antigen in the context of HLA-E providing evidence for a relevant role of HLA-E restricted CD8(+) T cells.In addition, our data indicate that HLA-E allelic variants may affect the natural course of HCV infection.

We compared lumbar and femoral bone density in 129 black (M age=61.0 +/- 12.7 yrs) and 130 white (M age 62.7 +/- 8.4 yrs) postmenopausal women volunteers. A total of 44/129 (34%) black Ss and 84/130 (65%) of white Ss were currently taking hormone replacement therapy (HRT) as prescribed by their personal physicians. Standing height and body weight were obtained on Detecto scales. We measured the bone tissue with a Hologic QDR-2000 densitometer, and compared the bone density for the L2-L4 lumbar spine (SP), and the total femur(HIP) sites, adjusting for years past menopause (YPM: yrs) and body mass index(BMI: wt(kg)/ht(m2)]. The following means and adjusted means were obtained: This table shows that, after adjusting for body mass index and years past menopause, the overall L2-L4 regional differences appear to be less clinically striking. The adjusted racial differences in the total hip, although a bit more pronounced, are not statistically significant. Additional analyses, however, did yield some significant differences at various subsites (data not shown). We believe that the consideration of weight-bearing hypotheses and other lifestyle and dietary factors could possibly erode the often apparently large racial differences, suggesting that blacks with these identified lifestyle factors (or their correlates) may have similar risks for osteoporotic fractures as their white counterparts.

This study compared femoral bone sites between two racial groups of similarly aged postmenopausal women. One hundred twenty nine black women(age=61.0±12.7 yrs) and 130 white women (age=62.7±8.4 yrs) volunteered for this study. Height and weight were obtained and a questionnaire was given orally to assess activity, gynecological, medical, and nutritional histories. Bone measurements (Bone Mineral Content, BMC: Bone Area, BA: and Bone Mineral Density, BMD) were obtained at 5 femoral sites(Femoral Neck, FN; Trochanter, T; Inter Trochanter, IT; Ward's Triangle, WT; and Total Femur, TF) with Hologic QDR 2000 Bone Densitometer. Two-way ANOVAs(2 races by 4 age groups, <50, 50-59, 60-69, 70+ yrs) were used to determine differences in femoral bone between races, age groups, and for race by age interactions. The interactions compared bone change across age groups between the two races. Significant (p<.05) results revealed: 1) white women were taller (162.7 ±5.6 to 160.52±5.6 cm) but black women weighed more (83.3±21.2 to 65.0±12.3 kg), 2) black women had greater BMC and BMD values at all 5 femoral sites but no difference in BA, except at the T and TF sites where white women had larger BAs(10.876±1.458 to 10.402±1.414 and 34.467±3.197 to 33.326±3.872 cm2, respectively), 3) losses in BMD occurred across age groups in both races, and 4) no significant race by age interactions. It was concluded that black women had higher femoral BMC and BMD values but both races lost bone at similar rates. Thus, black postmenopausal women become at risk for osteoporotic fractures at an older age than white women.

This study compared anthropometric measurements between two racial groups of similarly aged postmenopausal women. One hundred twenty nine black women(age=61.0±12.7 yrs) and 130 white women (age=62.7±8.4 yrs) volunteered for this study. Standing and sitting heights, body weight, 4 skinfolds, and 4 body girths were obtained on both racial groups. Body Mass Index [BMI wt (kg)/ht (m2)] and Waist to Hip Ratio (WHR) were also calculated. Skinfolds (tricep. Tr; chest, Ch; midaxillary, Ma; and abdomen, Ab) were measured serially with Harpend en Calipers and body girths (forearm, chest, waist, gluteal) with a Gulick Tape according to Lohman. et al. (1980). Peak rt and l f t hand grips were measured with a dynamometer. Significant (p<.05)“t” tests between the two racial groups revealed: 1) white subjects (Ss) had taller mean sitting and standing heights than black Ss(86.7±2.9 to 82.6±3.4 cm and 162.5±5.6 to 160.5±5.6 cm. respectively) but black Ss weighed more (83.3±21.2 to 65.0±12.3 kg, respectively); 2) black Ss had larger values for BMI(32.3±8.1 to 24.5±4.3 kg/m2). WHR (.84±.08 to.78±.06), skinfolds (Tr=29.1±10.3 to 25.3±6.8 mm; Ch=21.8 ±8.9 to 14.5±5.8 mm; Ma=27.0±11.1 to 18.4±7.8 mm and Ab=43.7±13.2 to 29.8±8.9 mm, respectively) and 4 body girths; and 3) white Ss had greater peak rt and lft hand grip strengths (26.7±4.9 to 22.3±9.1 kg and 23.6±19.8 kg to 19.8±7.5 kg, respectively) than black Ss. It was concluded that the high level of body fat overall and the concentration of it in the mid area of the body put the black Ss at risk for diabetes, hypertension, and other diseases associated with obesity.

This study compared lean, fat, and bone tissue masses between two racial groups of similarly aged postmenopausal women. One hundred twenty nine black women (M age=61.0±12.7 yrs) and 130 white women (M age=62.7±8.4 yrs) volunteered for this study. Standing height and body weight were obtained on Detecto scales. Measurement of the three body tissues was made with an Hologic QDR 2000 Bone Densitometer. Tissue assessments were subdivided into 6 body segments and combined for the total body. Significant (p<.05)“t” tests, used to determine racial differences in height, weight and tissue masses for 6 body segments and for total body, revealed: 1) white subjects (Ss) were taller (162.7 ±5.6 to 160.52±5.6 cm) but black Ss weighed more (83.3±21.2 to 65.0±12.3 kg), 2) black Ss had greater lean, fat, and bone tissue masses than white Ss at each body segment and for the total body (M total lean=43.4±6.7 to 37.1±4.2 kg; M total fat=38.7± 15.7 to 26.9±9.8 kg; and M total bone=2.3±0.4 to 2.0±0.3 kg, respectively). Significant(p<.05) one-way ANOVAS, used in each racial group across 4 cross-sectional age groups, <50, 50-59, 60-69, 70+ yrs, revealed black and white Ss lost significant lean and bone tissue masses (9.9% to 8.1% and 16.1% to 17.5%, respectively) between youngest and oldest groups but only white Ss lost fat tissue mass across age groups. It was concluded in this study, that black Ss had greater lean, fat, and bone mass tissues in 6 body segments and total body than white Ss. However, both racial groups lost lean and bone tissues similarly while only white Ss lost fat tissue across age groups.

This study compared radial and spinal bone sites between two racial groups of similarly aged postmenopausal women. One hundred twenty nine black women(age=61.0±12.7 yrs) and 130 white women (age=62.7±8.4 yrs) volunteered for this study. Height and weight were obtained on Detecto Scales. Bone measurements (Bone Mineral Content, BMC; Bone Area, BA; and Bone Mineral Density, BMD) were obtained at 3 radial sites and at the mean of Anterior/Posterior Lumbar2-4 (L2-4) with Hologic QDR 2000 Bone Densitometer. Significant (p<.05) “t” tests results revealed: 1) white subjects (Ss) were taller (162.7 ±5.6 to 160.52±5.6 cm, respectively) but black Ss weighed more (83.3±21.2 to 65.0±12.3 kg, respectively); and 2) black Ss greater mean BMD values than white Ss at 3 radial sites (1/3 BMD=0.665±0.085 to 0.618±.076 g/cm2; Mid BMD=0.584±.077 to 0.543±0.068 g/cm2; and Ultra Distal BMD=.436±.077 to 0.400±0.066 g/cm2, respectively) at spinal site (L2-4=1.051±0.164 to.922±0.154 g/cm2, respectively). Significant (p<.05) one-way ANOVAs in each race across 4 age groups (<50, 50-59, 60-69, and 70+ yrs) revealed: 1) losses in BMD in both races, and 2) percentage loss of BMD slightly greater for white Ss than for black Ss at radial sites (1/3=15% to 13%; Mid=16% to 13%; and Ultra Distal=17% to 17%) but much greater loss at spinal site (L2-4=16% to 9%). It was concluded that black Ss had higher radial and spinal BMD values and lost bone similarly in the forearm but much slower in the spine than white Ss. Thus, black postmenopausal women become at risk for osteoporotic fractures of radius or spine at an older age than white women.

We evaluated body composition parameters in 71 postmenopausal (PM) women (X age=67.4±4.9 yrs on 10 yr test) followed for 10 yrs. Height, weight, skinfold measurements at 8 sites (tricep, subscapular, chest, mid-axillary, suprailiac, abdomen, thigh, calf), body girths at 7 sites (relaxed bicep, contracted bicep, forearm, chest, waist, thigh, and calf), and radial bone measurement (Bone Mineral Content, BMC, g; Bone Width, BW, cm; and BMC/BW g/cm2) at 1/3 and 4 mm distal sites were assessed with a Norland SPA Bone Densitometer on a pretest, 5-yr test, and a 10-yr test. One technician did all bone testing. One-way ANOVAs, used to determine changes across time in these PM women, revealed at the.05 level: 1) Significant decreases in mean standing height between pretest and 5-yr test and between 5 and 10 yr tests(162.1±4.9; 161.3±4.9 and 160.5±5.0 cm, respectively); 2) No significant change in body weight; 3) Significant mean increases in skinfold thicknesses between pretest and 10 yr-test for tricep(20.1±5.5 to 21.0±6.0 mm), subscapular (15.3±5.9 to 17.7±8.2 mm), suprailiac (16.6±8.4 to 18.1±8.4 mm), abdomen(29.3±10.5 to 32.2±11.5 mm), & calf (19.2±5.6 to 21.9±7.5 mm) skinfolds; 4) Significant mean increases between pretest and 10 yr test in chest (88.1±6.5 to 93.5±8.1 cm), waist(74.1±8.8 to 81.1±10.8 cm), thigh (46.0±3.9 to 49.5±4.4 cm), and calf (34.4±2.5 to 35.7±2.8 cm) circumferences; 5) Significant decreases in BMC and BMC/BW at the 1/3 distal radial sites (0.784±111 to 0.781±.128 g, and 0.667±0.076 to 0.657±0.091 g/cm2, respectively); and 6) No significant changes in bone mass at the 4 mm site. On the basis of these data, it was concluded, that as PM women aged, fat tissue mass increased primarily in trunk area without concomitant change in body weight.

We evaluated the effects of exercise (EXC) and estrogen replacement therapy(ERT) upon radial bone mass in 71 postmenopausal (PM) women followed for 10 yrs. Multiple regression (REG) equations predicted future bone levels in individual Ss under hypothetical interventions of all combinations of EXC and/or ERT. Radial bone mass at the 1/3 and 4mm sites were measured in 1984(BL=baseline), 1989 (BL+5), and 1994 (BL+10) using a Norland SPA Bone densitometer. EXC consisted of light weight lifting (40-60% of 1 RM for benchpress, military press, reverse fly, lat pull), heavy hands (1-5 lbs) during supervised aerobic exercise and walking. ERT consisted primarily of Premarin usage (0.625 mg). REG analyses showed ERT usage to be a significant predictor (p<0.05) of future bone mass at both sites at BL+5 and BL+10(adjusting for individual covariates such as years past menopause and pre-baseline history of EXC and/or ERT usage). EXC was a significant predictor of bone mass at both sites for BL+5, but not for BL+10. R2 values were 0.90 (1/3,BL+5), 0.92 (4mm,BL+5), 0.78 (1/3,BL+10), and 0.85 (4mm,BL+10). PM women should be educated regarding the usefulness of combinations of ERT and EXC as interventions for preserving bone mass. More importantly, the high R2 values suggest the equations can be used clinically for efficient individual predictions of bone levels that may indicate whether intervention(s) are warranted, depending on comparisons with known fracture thresholds.