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A novel diagnostic immunoassay testing procedure for hepatitis B virus core antibody (anti-HBc) using homogeneous purified full-length hepatitis B virus core antigen (HBcAg) capsids obtained from Escherichia coli was compared with Abbott Architect anti-HBc chemiluminescent microparticle immunoassay (CMIA; indirect method) against a library of specimens. A monoclonal anti-HBc neutralization confirmatory assay was then used to determine the degree of discordance between specimens. The new assay was found to be superior in both sensitivity and specificity.Antibodies (anti-HBc) to hepatitis B virus (HBV) core antigen (HBcAg) are present in current and past HBV infections. Anti-HBc antibodies represent a long-term serological marker of HBV infection, initially appearing during the acute phase of the infection and generally persisting thereafter (16). Thus, as anti-HBc is a universal marker of HBV infection, routine blood donor screening for anti-HBc has been implemented in some countries with low endemicity. It is a cost-effective method of screening. Such screening procedures have resulted in a decrease in the risk of posttransfusion HBV infections (12). In some individuals, the only serological marker of HBV infection is the presence of anti-HBc antibodies (10, 24), and thus detection of "anti-HBcAg alone" could reflect unrecognized "occult" HBV infection and physicians should consider investigating such patients with HBV molecular tests (21). Additionally, isolated anti-HBc can be used as a marker to assess the risk of HBV reactivation in patients undergoing therapy that could result in immunosuppression or patients who are HIV positive (17) or hepatitis C virus (HCV) positive (25).With these uses in mind, having a more efficient and reliable assay for anti-HBc is desirable. Most current commercially available anti-HBc assays have poor sensitivity or specificity (2, 19) and can be attributed to the inferior performance of the competitive immunoassay, especially for detecting low-titer antiHBc-reactive samples. False-positive reactivity can partially be attributed to unspecific activation of premature B lymphocytes causing the production of IgM, IgA, or IgM-related molecules without previous exposure to HBV (18,19). The specificity of competitive assays for anti-HBc can be significantly improved by addition of mild reducing agents, but such modified procedures often lead to the loss of sensitivity, particularly for IgM anti-HBc (23). In this study, a novel immunoassay for anti-HBc based on the double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) method is compared with a commercial anti-HBc assay, the Architect chemiluminescent microparticle immunoassay (CMIA). MATERIALS AND METHODSExpression and purification of full-length rHBcAg in E. coli. Two full-length HBcAg genes were obtained by chemosynthesis: AB090269 (genotype D, named CpD) and AB014368 (genotype C, named CpC). The rare arginine codes in Escherichia coli (AGA and AGG) located in the HBcAg gene were changed to CGT. The two fragments were inse...

BackgroundCDCA5 plays an important role in the development of various human cancers, but the associated mechanisms have not been investigated in hepatocellular carcinoma (HCC).Materials and methodsWe evaluated expression levels and functions of CDCA5 in HCC and showed that CDCA5 is upregulated in HCC tissues compared with paired or unpaired normal liver tissues.ResultsIncreased CDCA5 expression in HCCs was significantly associated with shorter survival of patients. Knockdown of CDCA5 using lentivirus-mediated shRNA significantly inhibited cell proliferation and suppressed cell survival, as well as induced cell cycle arrest at the G2/M phase and cell apoptosis of HCC cells. The tumor suppression effects of CDCA5 knockdown were mediated by decreased expression of cyclin-dependent kinase 1 (CDK1) and CyclinB1, which were increased in HCC tissues comparing with adjacent normal liver tissues. Moreover, upregulation of CDCA5 was positively associated with increased CDK1 and CyclinB1 expression in HCC tissues.ConclusionThe present data warrant consideration of CDCA5 as a prognostic biomarker and therapeutic target for HCC.

B- and T-lymphocyte attenuator (BTLA) is an immune-regulatory receptor, similar to CTLA-4 and PD-1, and is mainly expressed on B-, T-, and all mature lymphocyte cells. Herpes virus entry mediator (HVEM)-BTLA plays a critical role in immune tolerance and immune responses which are areas of intense research. However, the mechanisms of the BTLA and the BTLA/HVEM signaling pathway in human diseases remain unclear. This review describes the research milestones of BTLA and HVEM in chronological order and their role in chronic HBV infection.

Self-replicating (replicon) RNA is a promising new platform for gene therapy, but applications are still limited by short persistence of expression in most cell types and low levels of transgene expression in vivo . To address these shortcomings, we developed an in vitro evolution strategy and identified six mutations in nonstructural proteins (nsPs) of Venezuelan equine encephalitis (VEE) replicon that promoted subgenome expression in cells. Two mutations in nsP2 and nsP3 enhanced transgene expression, while three mutations in nsP3 regulated this expression. Replicons containing the most effective mutation combinations showed enhanced duration and cargo gene expression in vivo . In comparison to wildtype replicon, mutants expressing IL-2 injected into murine B16F10 melanoma showed 5.5-fold increase in intratumoral IL-2 and 2.1-fold increase in infiltrating CD8 T cells, resulting in significantly slowed tumor growth. Thus, these mutant replicons may be useful for improving RNA therapeutics for vaccination, cancer immunotherapy, and gene therapy.

Background and Aim The aim of this study is to evaluate the association of the regulatory T cells (Treg)/T‐helper (Th) 17 cells and transforming growth factor‐β1 (TGF‐β1)/interleukin‐17 (IL‐17) ratios with the survival and disease progression in patients with hepatitis B virus (HBV)‐associated liver cirrhosis (LC). Methods The frequencies of Treg and Th17 cells were analyzed in 28 patients with HBV‐LC, 70 patients with chronic hepatitis B (CHB) and 20 normal controls (NC) by flow cytometry. The levels of cytokines related to Treg/Th17 differentiation, including IL‐10, TGF‐β1, IL‐17, and IL‐23, were measured by ELISA. Results Compared with NC, Treg cells were significantly increased in CHB patients and slightly increased in HBV‐LC patients, whereas Th17 cells were markedly increased both in patients with CHB and HBV‐LC. HBV‐LC patients, especially the nonsurvival ones, manifested a profound decrease in the Treg/Th17 ratio, which was negatively correlated with Child‐Pugh and model of end‐stage liver disease scores. Serum IL‐10, TGF‐β1, IL‐17, and IL‐23 levels were all significantly higher in HBV‐LC patients than in NC. In addition, the TGF‐β1/IL‐17 ratio was also markedly increased in patients with HBV‐LC, especially in nonsurvival and decompensated liver cirrhosis patients, and positively correlated with total bilirubin, Child‐Pugh, and model of end‐stage liver disease scores. Conclusions The decreased Treg/Th17 ratio and increased TGF‐β1/IL‐17 ratio may be associated with the survival and disease progression in HBV‐LC patients, and both of the two ratios can be used independently to predict the prognosis and disease progression of HBV‐LC patients.

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The proinflammatory cytokines transforming growth factor beta 1 (TGF-β1) and interleukin (IL)-31 have been implicated in tissue injury. However, whether TGF-β1/IL-31 are stimulated and elevated in response to liver injury that leads to fibrogenesis in hepatitis B virus-related liver cirrhosis (HBV-LC) remains unclear. To investigate the association between TGF-β1/IL-31 and stages of chronic HBV infection, serum TGF-β1, IL-9, IL-10,IL-17, IL-22, IL-23, IL-31, IL-33, and IL-35 were determined among patients with chronic hepatitis B (CHB; n=19), HBV-LC (n=20), and a normal control population (NC; n=18). Disease severity in patients with HBV-LC was assessed using model for end-stage liver disease (MELD) scores. Serum TGF-β1 and IL-31 levels were strongly positively linked in all subjects, and both correlated positively with IL-22, IL-33, and IL-17. TGF-β1 and IL-31 levels in the blood were both significantly higher in CHB and HBV-LC patients than in NC subjects. Elevated serum TGF-β1 and IL-31 levels were positively associated with albumin, alpha-fetoprotein, creatinine, white blood cell count, and platelet levels. Serum TGF-β1 and IL-31 were markedly higher in HBV-LC patients who did not have esophageal varices, and IL-31 had the highest sensitivity and specificity (90.9% and 66.7%, respectively) for indicating the absence of this complication. In summary, TGF-β1 and IL-31 were linked to progression from CHB to LC, and correlated well with the severity of HBV-LC. These findings suggest possible roles of the TGF-β1/IL-31 pathway in the pathogenesis of liver fibrosis during chronic HBV infection.