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Human immunodeficiency virus (HIV)-infected patients must remain on a lifelong regimen of highly active antiretroviral therapy (HAART). The presence of a latent viral reservoir in resting CD4+ T cells necessitates long-term treatment to prevent HIV reactivation and spread from perpetuating disease. Latent reservoirs are therefore the main barrier to eradication of HIV infection. The challenges of long-term HAART treatment include limited penetration to tissues that harbor latent virus (such as the brain), prohibitive cost, and both short- and long-term toxic effects [1–4]. Minocycline, a second-generation tetracycline derivative, has a number of properties that address these concerns (enhanced penetration to tissues, low incidence of toxicity with long-term administration, low cost). Because of these properties, along with its immunomodulatory abilities, it may prove to be an effective complement to HAART [5, 6]. We have shown elsewhere that minocycline has multiple beneficial effects in our simian immunodeficiency virus (SIV) macaque model of HIV-associated neurological disease [7]. In the central nervous system (CNS), treatment with minocycline significantly decreased virus load in the cerebrospinal fluid, viral RNA in the brain, and the severity of CNS disease. In the periphery, plasma viral load was decreased in minocycline-treated animals with encephalitis compared with those without encephalitis. Minocycline also decreased p38 activation and HIV replication in primary human lymphocytes, which suggests that it acts through its effects on CD4+ T cells, the primary host cell for HIV infection. Minocycline has also been shown to affect T cell activation and proliferation. Synovial T cell clones derived from arthritic patients treated with minocycline in vitro possess an impaired proliferative response to anti-CD3 and decreased production of activation-induced cytokines [8]. The effect of minocycline was also found to be signal dependent, resulting in decreased levels of tumor necrosis factor (TNF)-α messenger RNA in response to stimulation with phorbol 12-myristate 13-acetate or anti-CD28 [9]. These in vitro observations, along with our in vivo findings in the SIV primate model, represent evidence that minocycline can modulate cellular activation and proliferation by altering cell signaling. Owing to the tightly coupled regulation of the HIV long terminal repeat (LTR) by many T cell regulatory signals and the dependence of productive HIV replication on the activation state of the host cell, we hypothesized that minocycline treatment would decrease HIV replication by decreasing the activation state of the host cell [10–12]. In this study, we demonstrate that minocycline decreases single-cycle HIV replication, intracellular viral RNA levels after in vitro infection of primary human CD4+ T cells, reactivation of HIV from a primary CD4+ T cell-derived model of HIV latency, and reactivation of HIV from resting CD4+ T cell reservoirs from patients who have clinically undetectable viremia during HAART. Furthermore, we demonstrate that minocycline alters T cell activation, blunting changes in expression of T cell activation/ proliferation markers and cytokine secretion, many of which are critical for activation pathways that regulate HIV replication.

This randomized, double-blind, placebo-controlled study evaluated whether lamivudine given during late pregnancy can reduce hepatitis B virus (HBV) perinatal transmission in highly viraemic mothers. Mothers were randomized to either lamivudine 100 mg or placebo from week 32 of gestation to week 4 postpartum. At birth, infants received recombinant HBV vaccine with or without HBIg and were followed until week 52. One hundred and fifty mothers, with a gestational age of 26-30 weeks and serum HBV DNA >1000 MEq/mL (bDNA assay), were treated. A total of 141 infants received immunoprophylaxis at birth. In lamivudine-treated mothers, 56 infants received vaccine + HBIg (lamivudine + vaccine + HBIg) and 26 infants received vaccine (lamivudine + vaccine). In placebo-treated mothers, 59 infants received vaccine + HBIg (placebo + vaccine + HBIg). At week 52, in the primary analyses where missing data was counted as failures, infants in the lamivudine + vaccine + HBIg group had a significant decrease in incidence of HBsAg seropositivity (10/56, 18%vs 23/59, 39%; P = 0.014) and in detectable HBV DNA (11/56, 20%vs 27/59, 46%; P = 0.003) compared to infants in the placebo + vaccine + HBIg group. Sensitivity analyses to evaluate the impact of missing data at week 52 resulting from a high dropout rate (13% in the lamivudine + vaccine + HBIg group and 31% in the placebo + vaccine + HBIg group) remained consistent with the primary analysis in that lower transmission rates were still observed in the infants of lamivudine-treated mothers, but the differences were not statistically significant. No safety concerns were noted in the lamivudine-treated mothers or their infants. Results of this study suggest that lamivudine reduced HBV transmission from highly viraemic mothers to their infants who received passive/active immunization.

CD4 + T-cell depletion is the hallmark of AIDS pathogenesis. Multiple mechanisms may contribute to the death of productively infected CD4 + T cells and innocent-bystander cells. In this study, we characterize a novel mechanism in which human immunodeficiency virus type 1 (HIV-1) infection preferentially depletes peripheral memory CD4 + T cells before the completion of reverse transcription. Using a recombinant HIV-1 carrying the green fluorescent protein reporter gene, we demonstrate that memory CD4 + T cells were susceptible to infection-induced cell death at a low multiplicity of infection. Infected memory CD4 + T cells underwent rapid necrotic cell death. Killing of host cells was dependent on X4 envelope-mediated viral fusion, but not on virion-associated Vpr or Nef. In contrast to peripheral resting CD4 + T cells, CD4 + T cells stimulated by mitogen or certain cytokines were resistant to HIV-1-induced early cell death. These results demonstrate that early steps in HIV-1 infection have a detrimental effect on certain subsets of CD4 + T cells. The early cell death may serve as a selective disadvantage for X4-tropic HIV-1 in acute infection but may play a role in accelerated disease progression, which is associated with the emergence of X4-tropic HIV-1 in the late stage of AIDS.

Elite suppressors maintain normal CD4 + T cell counts and viral loads of

Although combination therapy allows the suppression of human immunodeficiency virus type 1 (HIV-1) viremia to undetectable levels, eradication has not been achieved because the virus persists in cellular reservoirs, particularly the latent reservoir in resting CD4+T lymphocytes. We previously established a simian immunodeficiency virus (SIV)/macaque model to study latency. We describe here a novel mechanism for the induction of SIV from latently infected resting CD4+T cells. Several human cell lines including CEMx174 and Epstein-Barr virus-transformed human B-lymphoblastoid cell lines mediated contact-dependent activation of resting macaque T cells and induction of latent SIV. Antibody-blocking assays showed that interactions between the costimulatory molecule CD2 and its ligand CD58 were involved, whereas soluble factors and interactions between T-cell receptors and major histocompatibility complex class II were not. Combinations of specific antibodies to CD2 also induced T-cell activation and virus induction in human resting CD4+T cells carrying latent HIV-1. This is the first demonstration that costimulatory signals can induce latent virus without the coengagement of the T-cell receptor, and this study might provide insights into potential pathways to target latent HIV-1.

Viral latency is a reversibly non-productive state of infection that allows some viruses to evade host immune responses. As a consequence of its tropism for activated CD4(+) T cells, HIV-1 can establish latent infection in resting memory CD4(+) T cells, which are generated when activated CD4(+) T cells return to a quiescent state. Latent HIV-1 persists as a stably integrated but transcriptionally silent provirus. In this state, the virus is unaffected by immune responses or antiretroviral drugs, and this latent reservoir in resting CD4(+) T cells is a major barrier to curing the infection. Unfortunately, there is no simple assay to measure the number of latently infected cells in a patient, nor is there an entirely representative in vitro model in which to explore the molecular mechanisms of latency. This Review will consider current approaches to the analysis of HIV-1 latency both in vivo and in vitro.

The impact of ribavirin (RBV) dosage on sustained virologic response (SVR) rates remains elusive in hepatitis C virus genotype 2 (HCV-2) rapid responders receiving 16 weeks of peginterferon (Peg-IFN) plus RBV. Treatment-naïve HCV-2 patients with rapid virologic response (RVR) received Peg-IFN alfa-2a 180 μg/week plus weight-based RBV (1,000 or 1,200 mg/day; cut-off body weight: 75 kg) for 6 weeks and then randomly received Peg-IFN alfa-2a 180 μg/week plus weight-based (1,000 or 1,200 mg/day; n = 247) or flat-dose (800 mg/day; n = 246) RBV for additional 10 weeks. The primary endpoint was SVR24. Patients receiving weight-based and flat-dose RBV therapies had comparable SVR24 rates (93.5% versus 91.9%, P = 0.49). The risk differences (RDs) of SVR24 receiving weight-based and flat-dose RBV arms were 7.1% [95% CI: 0.7% to 13.6%] in males and −5.8% [95% CI: −12.1% to 0.5%] in females (interaction P = 0.01). The SVR24 rate was higher in males receiving ≥13 mg/kg/day than those receiving P = 0.001). In conclusion, Peg-IFN alfa-2a plus weight-based or flat-dose RBV for 16 weeks provides comparable SVR24 rates in treatment-naïve HCV-2 rapid responders. However, males should receive weight-based RBV to achieve a high SVR24 rate.

Hepatitis C virus genotype 2 (HCV-2) slow responders poorly respond to 24 weeks of peginterferon (Peg-IFN) plus ribavirin (RBV). We evaluated the efficacy of extended 48-week regimen and the role of interleukin-28B (IL-28B) genotype in this clinical setting. Treatment-naïve HCV-2 patients not achieving rapid virologic response (RVR) by Peg-IFN alfa-2a 180 μg/week plus weight-based RBV (1,000–1,200 mg/day, cutoff body weight of 75 kg) were randomly assigned to receive a total duration of 48 (n = 94) or 24 (n = 93) weeks of therapy. The primary endpoint was sustained virologic response (SVR). Baseline patient characteristics to predict SVR were analyzed. Patients receiving 48 weeks of treatment had a greater SVR rate than those receiving 24 weeks of treatment (70.2% versus 46.2%, P = 0.001). Compared to patients treated for 24 weeks, the SVR rate in those treated for 48 weeks increased by 10.9% [95% CI: −5.9% to 27.7%] and 65.6% [95% CI: 44.5% to 86.7%] if they had IL-28B rs8099917 TT genotype and GT/GG genotype, respectively (interaction P = 0.002). In conclusion, 48-week treatment with Peg-IFN plus weight-based RBV provides a greater SVR rate than 24-week treatment in treatment-naïve HCV-2 patients with unfavorable IL-28B genotypes who fail to achieve RVR.

Therapeutic vaccines are considered to be able to reverse the dysfunctional immune state of chronic hepatitis B and thus hold the promise for HBV cure. Martin et al. developed a novel adenovirus-based therapeutic vaccine TG1050 and demonstrated its induction of long-lasting antiviral CD8+ T-cell immunity in mouse models of HBV persistence.

Neutralizing antibodies (NAb) against autologous virus can reach high titers in human immunodeficiency virus type 1 (HIV-1)-infected patients with progressive disease. Less is known about the role of NAb in HIV-1-infected patients with viral loads of env sequences from plasma viruses and proviruses in resting CD4 + T cells of HAART-treated patients, elite suppressors, and untreated HIV-1-infected patients with progressive disease. For each patient group, we assessed plasma virus neutralization by autologous, contemporaneous plasma. The degree of env diversity, the number of N-linked glycosylation sites, and the lengths of variable loops were all lower in elite suppressors than in HAART-treated and untreated viremic patients. Both elite suppressors and HAART-treated patients had lower titers of NAb against HIV-1 lab strains than those of untreated viremic patients. Surprisingly, titers of NAb against autologous, contemporaneous plasma viruses were similarly low in chronic progressors, elite suppressors, and HAART-treated patients. In elite suppressors and HAART-treated patients, titers of NAb against autologous plasma viruses also did not differ significantly from titers against autologous proviruses from resting CD4 + T cells. These results suggest that high-titer NAb are not required for maintenance of viral suppression in elite suppressors and that NAb do not select plasma virus variants in most HAART-treated patients. Both drug-mediated and natural suppression of HIV-1 replication to levels below 50 copies/ml may limit the stimulation and maintenance of effective NAb responses.

Background/aims The prevalence and clinical implications of non-alcoholic fatty liver disease in patients with chronic hepatitis C remain unknown in Taiwan. Methods We addressed the relevant issues by analysing 95 naive Taiwanese patients infected with either hepatitis C virus (HCV) genotype 1 ( n=57) or 2 ( n=38), receiving interferon alone ( n=41) or in combination with ribavirin ( n=54) therapy. A single pathologist scored steatosis and steatohepatitis at baseline and 24 weeks after antiviral treatment. Results At baseline, steatosis and steatohepatitis were present in 44 (46%) and four (4%) patients, respectively. Variables associated with steatosis in logistical regression were hyperglycaemia ( P=0.01), hypertriglyceridaemia ( P=004) and body mass index ≥27 ( P=0.009), but not HCV genotype or viral load. The grade of steatosis correlated well with the number of metabolic syndrome parameters ( P=0.018). Interferon monotherapy, advanced age and HCV genotype 1, but not steatosis, correlated with lower sustained response rate. After treatment, steatosis improved in 19 and worsened in nine, which also did not correlate with HCV genotype ( P=0.850) or sustained response to antiviral therapy ( P=0.246). Conclusions Hepatic steatosis in Taiwanese patients with chronic hepatitis C was associated with features of the metabolic syndrome, but did not correlate with HCV genotype, advanced fibrosis or the response to antiviral therapy.

The latent reservoir for HIV-1 in resting memory CD4+ T cells is the major barrier to curing HIV-1 infection. Studies of HIV-1 latency have focused on regulation of viral gene expression in cells in which latent infection is established. However, it remains unclear how infection initially becomes latent. Here we described a unique set of properties of CD4+ T cells undergoing effector-to-memory transition including temporary upregulation of CCR5 expression and rapid downregulation of cellular gene transcription. These cells allowed completion of steps in the HIV-1 life cycle through integration but suppressed HIV-1 gene transcription, thus allowing the establishment of latency. CD4+ T cells in this stage were substantially more permissive for HIV-1 latent infection than other CD4+ T cells. Establishment of latent HIV-1 infection in CD4+ T could be inhibited by viral-specific CD8+ T cells, a result with implications for elimination of latent HIV-1 infection by T cell-based vaccines.

T cell immunity targeting the conserved epitopes of influenza virus has the potential to provide cross-protection against distinctly-related strains. Peptide vaccines are an ideal strategy to induce specific antiviral T cell immunity, but subimmunogenic stimulation by peptide vaccines tend to induce regulatory T (Treg) cells. However, it remains unclear about the roles of vaccine-induced antigen-specific Treg cells in vivo. Here, we aimed to investigate how vaccine-induced antigen-specific Treg cells respond during acute influenza virus infection, and the effects of adjuvants on them. By adoptive transfer experiments with OT-II TCR transgenic T cells, we found that the OVA OT-II peptide vaccine induced Treg cells and secondary vaccination further expanded them. Infection with influenza virus containing the OVA OT-II peptide also drove the expansion of pre-existing vaccine-induced Treg cells. Importantly, specific depletion of vaccine-induced antigen-specific Treg cells enhanced the antiviral immunity. Vaccination combined with adjuvants, especially with CpG, suppressed the development of vaccine-induced antigen-specific Treg cells. Finally, CpG-adjuvanted OVA protein or whole inactivated influenza vaccine by the subcutaneous-prime-intranasal-boost strategy reduced the ratio of antigen-specific Treg cells in lung and protected mice from the heterosubtypic influenza infection. In conclusion, unadjuvanted peptide vaccines promoted antigen-specific Treg cells, which were further expanded upon acute influenza infection. The CpG adjuvant restricted the development of vaccine-induced antigen-specific Treg cells, and enhanced the antiviral T cell immunity against heterosubtypic influenza infection.

Precore (PC) variant (G1896A) and basal core promoter (BCP) variant (A1762T/G1764A) of HBV are associated with risk of hepatocellular carcinoma in HBV carriers. However, little is known about their impact on the adverse outcomes of hepatitis B e antigen (HBeAg)-negative hepatitis and liver cirrhosis.251 spontaneous HBeAg seroconverters who had genotype B or C infection and received a long-term follow-up were enrolled. PC and BCP mutants were determined qualitatively and quantitatively to correlate with these adverse outcomes. The findings were validated by an independent case-control study, which included 184 patients with biopsy-proven liver fibrosis stages.In the longitudinal cohort study, BCP mutant and possibly PC wild type were associated with cirrhosis development, but not HBeAg-negative hepatitis. Multivariable analysis showed that only BCP mutant was an independent risk factor for cirrhosis development. Using quantitative analysis of BCP mutant, a higher proportion of BCP mutant, defined as a continuous variable, a dichotomous variable or an ordinal variable, was associated with a higher risk of cirrhosis. If we chose 45% of BCP mutant as the cut-off, the risk of cirrhosis was higher in patients with BCP mutant ≥45% compared to45% in the longitudinal cohort; this finding was validated by the case-control study (adjusted OR: 2.81, 95% CI 1.40 to 5.67).A higher proportion of BCP mutant increases the risk of liver cirrhosis development in HBV carriers with genotype B or C infection.

Data comparing the efficacy and safety of combination therapy with peginterferon plus low-dose ribavirin and peginterferon monotherapy in treatment-naive haemodialysis patients with hepatitis C virus genotype 2 (HCV-2) infection are limited.In this randomised trial, 172 patients received 24 weeks of peginterferon alfa-2a 135 μg/week plus ribavirin 200 mg/day (n=86) or peginterferon alfa-2a 135 μg/week (n=86). The efficacy and safety endpoints were sustained virological response (SVR) rate and adverse event (AE)-related withdrawal rate.Compared with monotherapy, combination therapy had a greater SVR rate (74% vs 44%, relative risk (RR): 1.68 [95% CI 1.29 to 2.20]; p0.001). The beneficial effect of combination therapy was more pronounced in patients with baseline viral load ≥800,000 IU/mL than those with baseline viral load800,000 IU/mL (RR: 3.08 [95% CI 1.80 to 5.29] vs. RR: 1.11 [95% CI 0.83 to 1.45]; interaction p=0.001). Patients receiving combination therapy were more likely to have a haemoglobin level of8.5 g/dL (70% vs. 8%, risk difference (RD): 62% [95% CI 50% to 73%]; p0.001) and required a higher dosage [mean: 13,417 vs. 6667 IU/week, p=0.027] of epoetin β to manage anaemia than those receiving monotherapy. The AE-related withdrawal rates were 6% and 3% in combination therapy and monotherapy groups, respectively (RD: 2% [95% CI -4% to 9%]).In treatment-naive haemodialysis patients with HCV-2 infection, combination therapy with peginterferon plus low-dose ribavirin achieved a greater SVR rate than peginterferon monotherapy. Most haemodialysis patients can tolerate combination therapy.ClinicalTrial.gov number, NCT00491244.

Acute exacerbation (AE) of chronic hepatitis B virus (HBV) infection is common and negatively impacts the clinical outcome. Although upsurge of viral load always precedes or coincides with AE, the underlying immunological mechanisms remain unclear and were investigated.We prospectively followed the serum cytokine/chemokine profiles, viral load, and alanine aminotransferase (ALT) levels in 250 patients and identified 44 consecutive patients (male: 72.7%; age: 40.4 ± 9.7 years; hepatitis B e antigen [HBeAg] positivity: 63.6%; genotype B/C: 75%/25%) who developed AE during the follow-up in a medical center. The impact of clinical characteristics (age, gender, HBeAg, ALT, HBV genotype), cytokines (tumor necrosis factor-alpha, interferon gamma, interleukin [IL]-2, IL-4, IL-6, and IL-10), and chemokines (CXCL10/interferon gamma-induced protein [IP]-10, CCL2/MCP-1, CXCL9/MIG, CCL5/RANTES, and CXCL8/IL-8) on the serum HBV DNA dynamics at different time points (baseline, peak of serum HBV DNA level, peak of serum ALT level, and after AE) were analyzed.Of 44 patients, serum HBV DNA level surged before the peak of serum ALT level in 23 (52.3%), and coincided with the peak of ALT in 21 (47.7%). The upsurge of serum viral load significantly correlated with the increase of IL-10 (P = 0.0037) and CXCL10/IP-10 (P = 0.0044). Upsurge of serum viral load was preceded by an increase in serum IL-4 (P0.05), IL-6 (P0.05), and IL-10 (P0.05). Combination of HBV genotype, IL-6 level at baseline, and ALT level at the peak of serum HBV DNA reliably predicted subsequent AE pattern (P = 0.0116).Enhanced Th2 activity is likely involved in the surge of HBV DNA level before hepatitis exacerbation.

Hepatitis B virus causes chronic infection in about 300 million of people worldwide and results in about one million deaths a year, because of virus-induced end-stage liver disease and liver cancer. Mechanisms from acute hepatitis B virus (HBV) infection progress into chronicity, repeated bouts of inflammations were reviewed, with emphasis on the unusual modest innate immune responses against the virus. Chronic inflammation paves the way for malignant transformation of the infected hepatocytes. Recent studies in human and animal liver cancer models identify major genetic alteration pathways leading to carcinogenesis. A better understanding of HBV persistence and carcinogenesis will provide new insights for more effective interventions to control chronic hepatitis B and its sequelae in the future.

Persistence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) under current antiviral therapy is a major barrier to eradication of chronic hepatitis B (CHB). Curing CHB will require novel strategies for specific disruption of cccDNA. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is a newly developed tool for site-specific cleavage of DNA targets directed by a synthetic guide RNA (gRNA) base-paired to the target DNA sequence. To examine whether this system can cleave HBV genomes, we designed eight gRNAs against HBV of genotype A. With the HBV-specific gRNAs, the CRISPR/Cas9 system significantly reduced the production of HBV core and surface proteins in Huh-7 cells transfected with an HBV-expression vector. Among eight screened gRNAs, two effective ones were identified. Interestingly, one gRNA targeting the conserved HBV sequence acted against different genotypes. Using a hydrodynamics-HBV persistence mouse model, we further demonstrated that this system could cleave the intrahepatic HBV genome-containing plasmid and facilitate its clearance in vivo, resulting in reduction of serum surface antigen levels. These data suggest that the CRISPR/Cas9 system could disrupt the HBV-expressing templates both in vitro and in vivo, indicating its potential in eradicating persistent HBV infection.

The level of hepatitis B surface antigen (HBsAg) has been shown to complement hepatitis B virus (HBV)-DNA level in predicting disease progression in hepatitis B e antigen (HBeAg)-negative patients, especially those with low viral loads. Whether this finding could be seen in spontaneous HBeAg seroconverters remains unclear.A retrospective cohort of 390 Taiwanese spontaneous HBeAg seroconverters with a mean follow-up period of 7.4 years was enrolled. The relationships between HBV-DNA/HBsAg levels and HBeAg-negative hepatitis/active viral replication (HBV-DNA level ≥ 2000 IU/mL) were investigated.In the overall cohort, serum HBV-DNA level served as a better predictor for HBeAg-negative hepatitis compared with HBsAg level. However, in those with HBV-DNA level2000 IU/mL, a higher HBsAg level was associated with a higher risk of HBeAg-negative hepatitis (P = 0.015). Multivariate analysis showed the hazard ratio of HBsAg level ≥ 1000 IU/mL versus1000 IU/mL was 4.1 (95% confidence interval: 1.3-13.6). When using the end-point of active viral replication, HBsAg ≥ 1000 IU/mL remained as an independent risk factor, with a hazard ratio of 2.5 (95% confidence interval: 1.1-5.9).In spontaneous HBeAg seroconverters with HBV-DNA level2000 IU/mL, HBsAg level ≥ 1000 IU/mL is associated with increased risks of HBeAg-negative hepatitis and active viral replication. Combining HBV-DNA2000 IU/mL and HBsAg level1000 IU/mL may be used to define minimal viral activity.

BACKGROUND Data are limited on the efficacy and safety of pegylated interferon plus ribavirin for patients with hepatitis C virus genotype 1 (HCV-1) receiving hemodialysis. OBJECTIVE To compare the efficacy and safety of combination therapy with pegylated interferon plus low-dose ribavirin and pegylated interferon monotherapy for treatment-naive patients with HCV-1 receiving hemodialysis. DESIGN Open-label, randomized, controlled trial. (ClinicalTrials.gov: NCT00491244). SETTING 8 centers in Taiwan. PATIENTS 205 treatment-naive patients with HCV-1 receiving hemodialysis. INTERVENTION 48 weeks of pegylated interferon-α2a, 135 µg weekly, plus ribavirin, 200 mg daily (n = 103), or pegylated interferon-α2a, 135 µg weekly (n = 102). MEASUREMENTS Sustained virologic response rate and adverse event-related withdrawal rate. RESULTS Compared with monotherapy, combination therapy had a greater sustained virologic response rate (64% vs. 33%; relative risk, 1.92 [95% CI, 1.41 to 2.62]; P < 0.001). More patients receiving combination therapy had hemoglobin levels less than 8.5 g/dL than those receiving monotherapy (72% vs. 6%; risk difference, 66% [CI, 56% to 76%]; P < 0.001). Patients receiving combination therapy required a higher dosage (mean, 13 946 IU per week [SD, 6449] vs. 5833 IU per week [SD, 1169]; P = 0.006) and longer duration (mean, 29 weeks [SD, 9] vs. 18 weeks [SD, 7]; P = 0.004) of epoetin-β than patients receiving monotherapy. The adverse event-related withdrawal rates were 7% in the combination therapy group and 4% in the monotherapy group (risk difference, 3% [CI, -3% to 9%]). LIMITATION Open-label trial; results may not be generalizable to patients on peritoneal dialysis. CONCLUSION In treatment-naive patients with HCV-1 receiving hemodialysis, combination therapy with pegylated interferon plus low-dose ribavirin achieved a greater sustained virologic response rate than pegylated interferon monotherapy. PRIMARY FUNDING SOURCE National Center of Excellence for Clinical Trial and Research.

The latent reservoir for HIV-1 in resting memory CD4+ T cells is a major barrier to eradication. In vitro models involving transformed cell lines have been used to search for small molecules that reactivate latent HIV-1. Histone deacetylase (HDAC) inhibitors can reverse HIV-1 latent infection. Most studies on HDAC inhibitors have been performed in cell line models that differ in important aspects from the resting CD4+ T cells that harbour latent HIV-1 in vivo. Therefore, we evaluated the potency and kinetics of HDAC inhibitors in a primary cell model of HIV-1 latency that involves resting CD4+ T cells.A green fluorescent protein (GFP)-expressing reporter virus NL4-3-Δ6-drGFP was used to generate latent infection in Bcl-2-transduced primary CD4+ T cells. Seventeen HDAC inhibitors were tested in this primary cell model. The effects of these HDAC inhibitors on the reactivation of latent HIV-1 were determined and compared with anti-CD3 and anti-CD28 co-stimulation.In Bcl-2-transduced primary CD4+ T cells, short-term treatment with HDAC inhibitors resulted in very limited reactivation of latent HIV-1, while prolonged treatment greatly enhanced drug efficacy. The effects of HDAC inhibitors in reactivating latent HIV-1 correlated with their inhibitory effects on class I HDACs. Importantly, HIV-1 reactivated by HDAC inhibitors can quickly re-establish latent infection upon drug removal.We identified unique features of HDAC inhibitor-induced reactivation of latent HIV-1 in primary CD4+ T cells. Our findings may be useful for the design of eradication trials.

Background. The serum hepatitis B virus (HBV) surface antigen (HBsAg) level can predict hepatocellular carcinoma (HCC) development in hepatitis B e antigen (HBeAg)–negative patients with an HBV DNA level of

The inflammasome/caspase-1 signaling pathway in immune cells plays a critical role in bacterial pathogenesis; however, the regulation of this pathway in the gastric epithelium duringHelicobacter pyloriinfection is yet to be elucidated. Here, we investigated the effect of catechins (CAs), sialic acid (SA), or combination of CA and SA (CASA) onH. pylori-induced caspase-1-mediated epithelial damage, as well asH. pyloricolonizationin vitro(AGS cells) andin vivo(BALB/c mice). Our results indicate that the activity of caspase-1 and the expression of its downstream substrate IL-1βwere upregulated inH. pylori-infected AGS cells. In addition, we observed increased oxidative stress, NADPH oxidase gp91phox, CD68, caspase-1/IL-1β, and apoptosis, but decreased autophagy, in the gastric mucosa ofH. pylori-infected mice. We have further demonstrated that treatment with CASA led to synergistic anti-H. pyloriactivity and was more effective than treatment with CA or SA alone. In particular, treatment with CASA for 10 days eradicatedH. pyloriinfection in up to 95% ofH. pylori-infected mice. Taken together, we suggest that the pathogenesis ofH. pyloriinvolves a gastric epithelial inflammasome/caspase-1 signaling pathway, and our results show that CASA was able to attenuate this pathway and effectively eradicateH. pyloriinfection.

BACKGROUND & AIMS: Quantitative HBsAg has been recognized to assist in the management of chronic hepatitis B virus (HBV) infection. However, its role in disease monitoring of HBeAg-negative patients remains unclear. We aimed to investigate the longitudinal HBsAg change in HBeAg-negative carriers with HBV genotype B or C infection. METHODS: This is a retrospective cohort study conducted in a university hospital. Treatment-naïve HBeAg-negative carriers followed for more than 3 years were recruited. Their hepatitis activities were categorized by longitudinal HBV-DNA levels into high viral-load (HVL: HBV-DNA >/ =2000 IU/mL persistently), low viral-load (LVL: HBV-DNA 1000, 100-999

Dear Sir, We read with great interest the article by Bayliss et al 1 e-published in August 2016. HBV is a DNA virus and viral mutants develop during the course of persistent infection because of the spontaneous error of viral reverse transcription. Some of the viral mutants will be selected and become the predominant strains under the pressure of host immunity if they bear better replication fitness. Two common HBV mutants, mutations in precore stop codon (PC) (G1896A) and basal core promoter (BCP) (A1762T/G1764A), have been shown to abolish or reduce the production of HBeAg, respectively, which contribute to disease progression.2 In previous reports, these viral variants were determined using population sequencing, which is a qualitative assay thus limits …

Supplemental Digital Content is available in the text, Hepatitis B virus (HBV) and hepatitis C virus (HCV) infections are 2 major causes of chronic viral hepatitis. It is still unclear how HCV coinfection affects HBV replication and clinical outcomes in HBV/HCV coinfected patients. We conducted a longitudinal study, which enrolled 111 patients with HBV/HCV coinfection and 111 propensity score-matched controls with HBV monoinfection. Both groups had comparable baseline age, sex, fibrosis stage, levels of HBV DNA, and HBV surface antigen (HBsAg). The HCV coinfection and other host/viral factors were correlated with various outcomes, including HBsAg loss and cirrhosis/hepatocellular carcinoma (HCC) development. After a 10-year follow-up, we found that HCV coinfection itself was not associated with HBsAg loss. However, coinfected patients with alanine aminotransferase (ALT) level >80 U/L had a higher chance of HBsAg loss than those with ALT level ≤80 U/L [hazard ratio (95% confidence interval): 4.41 (1.75–11.15)] or matched controls with HBV monoinfection [hazard ratio (95% confidence interval): 3.40 (1.54–7.50)]. Besides, both HCV coinfection and higher ALT levels were associated with higher HCC risks and the HCC risks remained even after HBsAg loss in HBV/HCV con-infected patient. HCV coinfection is not associated with HBsAg loss. A higher ALT level is a major determinant of HBsAg loss in patients with HBV/HCV coinfection. Both HCV coinfection and a higher ALT level were independent risk factors of HCC.

Patients dually infected with hepatitis C virus (HCV)/hepatitis B virus (HBV) have a higher risk of developing advanced liver disease or hepatocellular carcinoma compared with monoinfected patients. Yet, there is a similar rate of sustained virologic response (SVR) after peginterferon alfa-2a and ribavirin combination therapy in these patients compared with HCV-monoinfected patients and a high hepatitis B surface antigen (HBsAg) seroclearance rate. The durability of hepatitis C and B clearance in coinfected patients was investigated in a 5-year follow-up study. Patients with active HCV genotype 1, both HBV-coinfected (n = 97) and HBV-monoinfected (n = 110), underwent 48-week combination therapy with peginterferon alfa-2a plus ribavirin. In patients with active HCV genotype 2 or 3, both HBV-coinfected (n = 64) and monoinfected (n = 50) patients underwent 24-week combination therapy. A total of 295 (91.9%) patients completed treatment and 24 weeks posttreatment follow-up; 264 (89.5%) patients agreed to receive additional follow-up for up to 5 years after the end of treatment. After a median follow-up of 4.6 ± 1.0 years, six of the 232 patients achieving SVR developed HCV RNA reappearance, including five HCV genotype 1/HBV-coinfected patients and one HCV genotype 2/3-monoinfected patient. Subgenomic analysis of the HCV core gene indicated that five patients developed delayed recurrence of HCV infection. Overall, the cumulative recurrence rate of HCV infection was 2.3% (0.4%/year; 95% confidence interval [CI], 0.9%-5.5%). The cumulative HBsAg seroclearance rate was 30.0% (95% CI, 21.5%-42.0%); with 33.1% (95% CI, 21.8%-50.1%) in the 48-week combination therapy group and 24.3% (95% CI, 13.7%-42.9%) in the 24-week therapy group. Conclusion: Peginterferon alfa-2a and ribavirin therapy provides good HCV SVR durability and a high accumulative HBsAg seroclearance rate in patients who are coinfected with HCV and HBV. (HEPATOLOGY 2013;)

Precore (PC) (G1896A) and basal core promoter (BCP) (A1762T/G1764A) mutations of the hepatitis B virus (HBV) genome often emerge in chronic hepatitis B (CHB) patients. Their roles in hepatitis B e antigen (HBeAg) seroconversion induced by interferon (IFN) therapy remain controversial, partly because quantitative analysis for these mutants is lacking. This study aimed to develop a new assay to accurately quantify the PC and BCP mutant percentages and correlate their dynamic changes with IFN-induced HBeAg seroconversion in HBeAg-positive CHB patients. The PC and BCP mutant percentages were analyzed by polymerase chain reaction (PCR)-pyrosequencing. Our results showed that this quantitative assay for PC and BCP mutants achieved high accuracy (R2 > 0.99) within a range between 10% and 90% mutants. We examined dynamic changes of the PC and BCP mutant percentages following IFN treatment in 203 HBeAg-positive CHB patients. By multiple logistic regression analysis, we found that the chance of HBeAg seroconversion increased by 2.2% (odds ratio [OR] = 1.022, 95% confidence interval [CI]: 1.009-1.034, P = 0.001) and 2.3% (OR = 1.023, 95% CI: 1.010-1.037, P = 0.001) per 1% increase of the pretreatment PC and BCP mutant percentages, respectively, after adjustment for other predictors. However, only the pretreatment PC mutation percentage was significantly associated with HBeAg seroconversion with HBV DNA < 2,000 IU/mL (OR = 1.030, 95% CI: 1.014-1.047, P < 0.001). Furthermore, the mutant percentage of PC, but not BCP, in patients achieving HBeAg seroclearance with HBV DNA < 20,000 IU/mL increased significantly during IFN treatment (P = 0.039). Interestingly, patients with HBeAg seroconversion who had a high PC mutant percentage at the end of IFN treatment tended to exhibit high viremia after seroconversion. Conclusion: Quantitative analysis of PC and BCP mutants can predict IFN-induced HBeAg seroconversion and demonstrate their distinct evolution patterns during HBeAg seroconversion. (HEPATOLOGY 2013)

Chronic hepatitis B patients with high viral loads are at increased risk of cirrhosis and hepatocellular carcinoma (HCC). In patients with low viral loads, higher hepatitis B surface antigen (HBsAg) levels have been shown to predict HCC development. However, little is known about the difference in risk for other hepatitis B virus (HBV)-related adverse outcomes with varying HBsAg levels. A total of 1,068 Taiwanese hepatitis B e antigen (HBeAg)-negative HBV carriers with serum HBV DNA level

Loss of hepatitis B surface antigen (HBsAg) usually indicates the cure of hepatitis B virus (HBV) infection. In spontaneous hepatitis B e antigen (HBeAg) seroconverters, lower serum HBsAg and HBV DNA levels have been shown to be associated with HBsAg loss over time. However, little is known about their impacts on HBsAg loss in HBeAg-negative patients with limited viral replication. A total of 688 HBeAg-negative patients with baseline serum HBV DNA levels2000 IU/mL were enrolled in Taiwan. The relationships of HBsAg and HBV DNA levels with subsequent HBsAg loss were investigated. In a mean follow-up of 11.6 years, the average annual rate of HBsAg loss was 1.6%. Baseline HBsAg and HBV DNA levels were inversely associated with subsequent HBsAg loss. When compared to patients who had HBsAg levels1000 IU/mL, the rates of HBsAg loss were significantly higher in patients with HBsAg levels of 100-999, 10-99, and10 IU/mL, with hazard ratios of 2.5 (95% confidence interval [CI], 1.6-4.0), 2.8 (95% CI, 1.6-5.0), and 13.2 (95% CI, 8.1-21.5), respectively. Multivariate analysis showed that HBsAg level, but not HBV DNA, remained as an independent factor. The adjusted hazard ratio of HBsAg loss was 13.2 (95% CI, 7.8-22.1) for HBsAg level10 versus ≥ 1000 IU/mL. When compared to HBV DNA level by receiver operating characteristic curve analysis, HBsAg level served as a better predictor of both 5-year and 10-year HBsAg loss.In HBeAg-negative patients with HBV genotype B or C infection who have HBV DNA level2000 IU/mL, HBsAg level10 IU/mL is the strongest predictor of HBsAg loss.

To provide updated points of view regarding the recent development and application of in-vitro primary cell models of HIV latency and their potential application in future studies of HIV eradication.It has been challenging to develop a primary cell model of HIV latency that can authentically recapitulate the quiescent features of resting CD4+ T cells. Recently, several articles have described different approaches that can generate latently HIV-infected resting CD4+ T cells in vitro. Some of them further demonstrated that the primary cell models of HIV latency are suitable for biochemical studies and drug screening.Recent progress in primary cell models of HIV latency has facilitated research on the mechanisms of HIV latency. These models also serve as a platform for the discovery of drugs that can purge the latent reservoir for HIV in resting CD4+ T cells. These studies will allow deeper insight to the mechanisms governing HIV latency and may help identify some candidate compounds for use in therapeutic strategies for the eradication of latent HIV.

The Web abounds with dyadic data that keeps increasing by every single second. Previous work has repeatedly shown the usefulness of extracting the interaction structure inside dyadic data [21, 9, 8]. A commonly used tool in extracting the underlying structure is the matrix factorization, whose fame was further boosted in the Netflix challenge [26]. When we were trying to replicate the same success on real-world Web dyadic data, we were seriously challenged by the scalability of available tools. We therefore in this paper report our efforts on scaling up the nonnegative matrix factorization (NMF) technique. We show that by carefully partitioning the data and arranging the computations to maximize data locality and parallelism, factorizing a tens of millions by hundreds of millions matrix with billions of nonzero cells can be accomplished within tens of hours. This result effectively assures practitioners of the scalability of NMF on Web-scale dyadic data.

The development of highly active antiretroviral therapy (HAART) to treat individuals infected with HIV-1 has dramatically improved patient outcomes, but HAART still fails to cure the infection. The latent viral reservoir in resting CD4+ T cells is a major barrier to virus eradication. Elimination of this reservoir requires reactivation of the latent virus. However, strategies for reactivating HIV-1 through nonspecific T cell activation have clinically unacceptable toxicities. We describe here the development of what we believe to be a novel in vitro model of HIV-1 latency that we used to search for compounds that can reverse latency. Human primary CD4+ T cells were transduced with the prosurvival molecule Bcl-2, and the resulting cells were shown to recapitulate the quiescent state of resting CD4+ T cells in vivo. Using this model system, we screened small-molecule libraries and identified a compound that reactivated latent HIV-1 without inducing global T cell activation, 5-hydroxynaphthalene-1,4-dione (5HN). Unlike previously described latency-reversing agents, 5HN activated latent HIV-1 through ROS and NF-kappaB without affecting nuclear factor of activated T cells (NFAT) and PKC, demonstrating that TCR pathways can be dissected and utilized to purge latent virus. Our study expands the number of classes of latency-reversing therapeutics and demonstrates the utility of this in vitro model for finding strategies to eradicate HIV-1 infection.

HIV-1 latency in resting CD4 + T cells represents a major barrier to virus eradication in patients on highly active antiretroviral therapy (HAART). Eliminating the latent HIV-1 reservoir may require the reactivation of viral gene expression in latently infected cells. Most approaches for reactivating latent HIV-1 require nonspecific T cell activation, which has potential toxicity. To identify factors for reactivating latent HIV-1 without inducing global T cell activation, we performed a previously undescribed unbiased screen for genes that could activate transcription from the HIV-1 LTR in an NF-κB-independent manner, and isolated an alternatively spliced form of the transcription factor Ets-1, ΔVII-Ets-1. ΔVII-Ets-1 activated HIV-1 transcription through 2 conserved regions in the LTR, and reactivated latent HIV-1 in cells from patients on HAART without causing significant T cell activation. Our results highlight the therapeutic potential of cellular factors for the reactivation of latent HIV-1 and provide an efficient approach for their identification.

The search engines that index the World Wide Web today use access methods based primarily on scanning, sorting, hashing, and partitioning (SSHP) techniques. The MapReduce framework is a distinguished example. Unlike DBMS, this search engine infrastructure provides few general tools for indexing user datasets. In particular, it does not include order-preserving tree indexes, even though they might have been built using such indexing components. Thus, data processing on these infrastructures is linearly scalable at best, while index-based techniques can be logarithmically scalable. DBMS have been using indexes to improve performance, especially on low-selectivity queries and joins. Therefore, it is natural to incorporate indexing into search-engine infrastructure. Recently, we proposed an extension of MapReduce called Map-Reduce-Merge to efficiently join heterogeneous datasets and executes relational algebra operations. Its vision was to extend search engine infrastructure so as to permit generic relational operations, expanding the scope of analysis of search engine content. In this paper we advocate incorporating yet another database primitive, indexing, into search engine data processing. We explore ways to build tree indexes using Hadoop MapReduce. We also incorporate a new primitive, Traverse , into the Map-Reduce-Merge framework. It can efficiently traverse index files, select data partitions, and limit the number of input partitions for a follow-up step of map, reduce, or merge.

Elite suppressors (ES) are untreated human immunodeficiency virus type 1 (HIV-1)-infected individuals who maintain normal CD4+T-cell counts and control viremia to levels that are below the limit of detection of current assays. The mechanisms involved in long-term control of viremia have not been fully elucidated. CD4+CD25+regulatory T cells (Tregs) downmodulate chronic inflammation by suppressing the activation and proliferation of effector lymphocytes. We found that while Tregs were functional in ES and patients on highly active antiretroviral therapy (HAART), ES maintained high levels of Tregs in peripheral blood mononuclear cells whereas patients on HAART had evidence of Treg depletion. We also demonstrated that Tregs can serve as reservoirs for HIV-1 in vivo. These data suggest that both direct infection by HIV-1 and tissue redistribution are possible explanations for declining FoxP3+Tregs in progressive HIV-1 infection. Furthermore, the maintenance of Tregs may be one mechanism associated with the nonprogressive nature of HIV-1 infection in ES.

Elite suppressors (ES) are untreated human immunodeficiency virus type 1 (HIV-1)-infected patients who maintain viral loads of + T-cell response in the ES appears to have prevented viral replication and thus the evolution toward a more fit variant. In addition, CD8 + T cells in the ES have selected for a rare mutation in an immunodominant HLA-B*27-restricted Gag epitope, which also has a negative impact on fitness. The results strongly suggest that through direct and indirect mechanisms, CD8 + T cells in some ES control HIV-1 isolates are capable of causing profound immunosuppression.

Map-Reduce is a programming model that enables easy development of scalable parallel applications to process a vast amount of data on large clusters of commodity machines. Through a simple interface with two functions, map and reduce, this model facilitates parallel implementation of many real-world tasks such as data processing jobs for search engines and machine learning. However,this model does not directly support processing multiple related heterogeneous datasets. While processing relational data is a common need, this limitation causes difficulties and/or inefficiency when Map-Reduce is applied on relational operations like joins. We improve Map-Reduce into a new model called Map-Reduce-Merge. It adds to Map-Reduce a Merge phase that can efficiently merge data already partitioned and sorted (or hashed) by map and reduce modules. We also demonstrate that this new model can express relational algebra operators as well as implement several join algorithms.

The green noise of a Nd:GdVO4 laser intracavity-doubled by type-I crystal BIBO was demonstrated. The noise reduction by enforcing a single-mode operation by volumetric Bragg grating in a V-shaped cavity is first time reported.

Data mining models are often implemented as program code and stored in an ad hoc fashion. In this paper we describe a methodology for developing model bases that can be implemented with only an extended relational database. This method stores models as what we call lightweight functions, which are straightforward textual representations of function values. Many applications use models that admit this approach. Business applications in particular, which can have a very large number of specialcase rules or business logic, are suitable for development with lightweight functions. Financial and forecasting applications give another example. We argue that the Lightweight Model Base offers some advantages for these applications. Introduction of lightweight stored functions in relational databases is a way to integrate the software-engineering methodology of tabledriven programming. This methodology advocates storing functions and data in tables. The computing process is just a mechanical evaluation of "joining" data and function relations. It would make stored business logic transparent for understanding and maintenance as relational data. Table-driven programming has much in common with statistics and data mining, and is a natural framework for combining data mining with databases.

Huei-Ling Huang,1 Pei-Hsuan Lu,1 Hung-Chih Yang,1 Gi-Da Lee,1,2 Han-Ru Li,1 Kuo-Chih Liao1 1Graduate Institute of Biomedical Engineering, National Chung Hsing University, 2Department of Radiology, Taichung Veterans General Hospital, Taichung,Taiwan Abstract: The aim of this research is to provide proof of principle by applying the fiber-optic triggered release of photo-thermally responsive liposomes embedded with gold nanoparticles (AuNPs) using a 200 µm fiber with 65 mW and 532 nm excitation for topical release in vivo. The tunable delivery function can be paired with an apoptosis biosensor based on the same fiber-optic configuration for providing real-time evaluation of chemotherapy efficacy in vivo to perform as a personalized chemotherapy system. The pattern of topical release triggered by laser excitation conveyed through optical fibers was monitored by the increase in fluorescence resulting from the dilution of self-quenching (75 mM) fluorescein encapsulated in liposomes. In in vitro studies (in 37°C phosphate buffer saline), the AuNP-embedded liposomes showed a more efficient triggered release (74.53%±1.63% in 40 minutes) than traditional temperature-responsive liposomes without AuNPs (14.53%±3.17%) or AuNP-liposomes without excitation (21.92%±2.08%) by spectroscopic measurements. Using the mouse xenograft studies, we first demonstrated that the encapsulation of fluorescein in liposomes resulted in a more substantial content retention (81%) in the tumor than for free fluorophores (14%) at 120minutes after administration from in vivo fluorescence imaging. Furthermore, the preliminary results also suggested the tunable release capability of the system by demonstrating consecutive triggered releases with fiber-optic guided laser excitation. Keywords: fiber-optic guided excitation, light excitation triggered release, photo-thermal responsive liposome, gold nanoparticles, tunable release in vivo

Elite suppressors (ES) are untreated human immunodeficiency virus type 1 (HIV-1)-infected individuals who control viremia to levels below the limit of detection of current assays. The mechanisms involved in this control have not been fully elucidated. Several studies have demonstrated that some ES are infected with defective viruses, but it remains unclear whether others are infected with replication-competent HIV-1. To answer this question, we used a sensitive coculture assay in an attempt to isolate replication-competent virus from a cohort of 10 ES. We successfully cultured six replication-competent isolates from 4 of the 10 ES. The frequency of latently infected cells in these patients was more than a log lower than that seen in patients on highly active antiretroviral therapy with undetectable viral loads. Full-length sequencing of all six isolates revealed no large deletions in any of the genes. A few mutations and small insertions and deletions were found in some isolates, but phenotypic analysis of the affected genes suggested that their function remained intact. Furthermore, all six isolates replicated as well as standard laboratory strains in vitro. The results suggest that some ES are infected with HIV-1 isolates that are fully replication competent and that long-term immunologic control of replication-competent HIV-1 is possible.

The aim of this study was to determine the efficacy of using the sound velocity and tissue attenuation to clinically discriminate breast cancer from healthy tissues. The methods for reconstructing the sound-velocity and attenuation-coefficient distributions were previously proposed and tested on tissue-mimicking phantoms. The methods require only raw channel data acquired by a linear transducer array, and therefore can be implemented on existing clinical systems. In this paper, these methods are tested on clinical data. A total of 19 biopsy-proven cases, consisting of 5 carcinomas (CAs), 7 fibroadenomas (FAs), 6 adipose tissue (fat), and 1 oil cyst, were evaluated. A single imaging setup consisting of a 5-MHz, 128-channel linear array was used to simultaneously obtain B-mode image data, time-of-flight data, and attenuation data. The sound velocity and attenuation coefficient can be reconstructed inside and outside a region of interest manually selected in the B-mode image. To reduce distortion caused by tissue inhomogeneities, an optimal filter derived from pulse-echo data - with water replacing the breast tissue - is applied. We found that the sound velocities in CA, FA, and fat tissues relative to those in the surrounding tissues were 44.9plusmn35.9, 13.0plusmn40.5, and -55.9plusmn30.9 m/s (meanplusmnSD), respectively, whereas the relative attenuation coefficients were -0.30plusmn0.45, 0.24plusmn1.59, and 0.16plusmn0.64 dB/cm/MHz. These results indicate that CA can be discriminated from FA and fat by choosing an appropriate threshold for the relative sound velocity (i.e., 18.5 m/s). However, the large variations in the attenuation within the same type of tissue make simple thresholding ineffective. Nevertheless, the method described in this paper has the potential to reduce negative biopsies and to improve the accuracy of breast cancer detection in clinics

Hepatitis C virus (HCV) infection frequently persists despite substantial virus-specific cellular immune responses. To determine if immunologically driven sequence variation occurs with HCV persistence, we coordinately analyzed sequence evolution and CD8+ T cell responses to epitopes covering the entire HCV polyprotein in subjects who were followed prospectively from before infection to beyond the first year. There were no substitutions in T cell epitopes for a year after infection in a subject who cleared viremia. In contrast, in subjects with persistent viremia and detectable T cell responses, we observed substitutions in 69% of T cell epitopes, and every subject had a substitution in at least one epitope. In addition, amino acid substitutions occurred 13-fold more often within than outside T cell epitopes (P < 0.001, range 5–38). T lymphocyte recognition of 8 of 10 mutant peptides was markedly reduced compared with the initial sequence, indicating viral escape. Of 16 nonenvelope substitutions that occurred outside of known T cell epitopes, 8 represented conversion to consensus (P = 0.015). These findings reveal two distinct mechanisms of sequence evolution involved in HCV persistence: viral escape from CD8+ T cell responses and optimization of replicative capacity.