Avasimibe can cooperate with a DC-targeting and integration-deficient lentivector to induce stronger HBV specific T cytotoxic response by regulating cholesterol metabolism.

We have reported a lentivector which could effectively induce HBV-specific cytotoxic T lymphocytes (CTLs). Avasimibe is an inhibitor of acetyl-CoA acetyltransferase-1 (ACAT1), and has been shown to enhance T lymphocyte cytotoxicity on tumor cells. However, the role of avasimibe in lentivector-induced HBV-specific T cytotoxic response remains unknown. Based on previous study, we constructed an integration-deficient lentivector LVDC-ID-HBV (harboring HBcAg expression), and the in vitro experiments showed that the combination of avasimibe exhibited better efficacy in inducing HBV-specific CTL responses including cell proliferation, production of cytokines, as well as CTL killing activities. Mechanism experiments showed that increasing cell membrane cholesterol levels by MßCD-coated cholesterol or ACAT1 inhibition efficiently promoted TCR clustering, signaling transduction and immunological synapse formation, thereby mediating augmented CTL responses. Nevertheless, the depletion of plasma membrane cholesterol with MßCD led to obviously decreased CTL responses. The avasimibe-mediated strengthened immune effects were also determined in animal experiments and the results were in agreement with those from the in vitro research. In particular, the in vivo CTL killing activities were identified by the CFSE or BV-labeled splenocyte lysis assay. Moreover, the experiments in HBV transgenic mice showed that the LVDC-ID-HBV plus avasimibe group demonstrated the lowest serum HBsAg and HBV DNA levels, as well as the lowest expression of HBsAg and HBcAg in liver tissues. We concluded that the HBV-specific CTL immune responses could be potentiated by avasimibe through regulating plasma membrane cholesterol levels. Avasimibe may be a potential adjuvant for lentivector vaccine against HBV infection.

Exosomes loading Tapasin enhance T cell immune response by autophagy to inhibit HBV replication.

Hepatitis B virus (HBV) specific T cell immune response plays a vital role in viral clearance. Dendritic cell derived exosomes (Dexs) can activate T cell immunity effectively. Tapasin (TPN) is involved in antigen processing and specific immune recognition. In the present study, we elucidated that Dexs loading TPN (TPN-Dexs) could enhance CD8+ T cell immune response and inhibit virus replication in HBV transgenic mice. T cell immune response and the ability of inhibiting HBV replication were measured in HBV transgenic mice immunized with TPN-Dexs. Meanwhile, CD8+ T cell autophagy and specific T cell immune responses were measured in vitro and vivo, and the mechanisms probably involved in were explored. Purified TPN-Dexs could be taken up into the cytoplasm of DCs and upregulate CD8+ T cell autophagy to enhance specific T cell immune response. In addition, TPN-Dexs could increase the expression of AKT and decrease the expression of mTOR in CD8+ T cells. Further research confirmed that TPN-Dexs could inhibit virus replication and decrease the expression of HBsAg in the liver of HBV transgenic mice. Nevertheless, those also could elicit mice hepatocytes damage. In conclusion, TPN-Dexs could enhance specific CD8+ T cell immune responses via the AKT/mTOR pathway to regulate the autophagy and exert the antiviral effect in HBV transgenic mice.

Collagen XV mediated the epithelial-mesenchymal transition to inhibit hepatocellular carcinoma metastasis.

Background: Hepatocellular carcinoma (HCC) is a malignant cancer with rapid progression, vascular invasion, a high recurrence rate and poor prognosis, so it is necessary to take early measures to halt this process. Accumulating evidence indicates that collagen XV (translated by Col15a1) is a basement membrane molecule related to tumour metastasis in several organs. However, the potential function of collagen XV in the liver associated with HCC remains to be further elucidated. Methods: Col15a1 was overexpressed in HepG2 and HCCLM3 cells. CCK8 and colony formation assays were used to assess the capacity of cell proliferation, and Transwell and wound healing assays were utilized to measure cell migration. Western blotting and real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) quantified the protein and mRNA expression levels of genes related to the epithelial-mesenchymal transition (EMT). Then, the effect of collagen XV on tumour metastasis was confirmed in vivo. Finally, we inhibited discoidin domain receptor 1 (DDR1) via DDR1-IN-1 to explore whether the collagen XV interacted with DDR1 to regulate EMT. Results: Patients of HCC with higher expression of Col15a1 showed better survival than patients with low expression. Overexpression of collagen XV in HepG2 and HCCLM3 cells suppressed cell proliferation and migration in vitro and inhibited pulmonary and liver metastasis in vivo. In addition, collagen XV downregulated the DDR1 and transcription factor (Snail, Slug), regulated the EMT markers (Vimentin, E-cadherin, N-cadherin, and MMP9). Furthermore, inhibition of the DDR1 receptor by DDR1-IN-1 suppressed the gene promoting the EMT. Conclusions: Collagen XV functioned as a metastasis inhibitor in HCC by regulating the DDR1-Snail/Slug axis to regulate EMT.

Ubiquitinated Hepatitis D Antigen-Loaded Microvesicles Induce a Potent Specific Cellular Immune Response to Inhibit HDV Replication in Vivo.

Hepatitis D is the most severe form of human viral hepatitis and currently lacks an efficient therapy. Dendritic cell-derived exosomes (Dexs) have been found to induce immune responses capable of eliminating viruses. However, the therapeutic potential of antigen-loaded exosomes in hepatitis D is still unknown. Recently, we designed exosomes loaded with ubiquitinated hepatitis delta virus (HDV) small delta antigen (Ub-S-HDAg) and then treated mice bearing replicating HDV with these exosomes to explore their antiviral effect and mechanism. Mature dendritic cell-derived exosomes (mDexs) were loaded with Ub-S-HDAg and their antivirus function was evaluated in mice with HDV viremia. Furthermore, the proportion of CD8+ cells, the ratio of Th1/Th2 cells, the postimmunization levels of cytokines were explored, and the Janus kinases (JAK)/signal transducer and activator of transcription (STAT) pathway was evaluated with a JAK2 inhibitor AG490. In Ub-S-HDAg-Dexs group, the HDV RNA viral load was significantly decreased compared with other groups by CD8+ cell enrichment and an increase Th1/Th2 cell ratio. Furthermore, lymphocyte infiltration was increased, while the HDAg level was decreased in mouse liver tissue. However, there were no significant differences in HBV surface antigen (HBsAg), alanine aminotransferase (ALT), or aspartate aminotransferase (AST) levels among the groups. Moreover, p-JAK2, p-STAT1, p-STAT4, STAT1, and STAT4 expression was increased in Ub-S-HDAg-Dexs group. In conclusion, Ub-S-HDAg-Dexs might be a potential immunotherapeutic agent for eradicating HDV by inducing specific cellular immune response via the JAK/STAT pathway. IMPORTANCE Hepatitis D is the most severe viral hepatitis with accelerating the process of liver cirrhosis and increasing the risk of hepatocellular carcinoma. However, there are no effective antiviral drugs. Exosomes derived from mature dendritic cells are used not only as immunomodulators, but also as biological carriers to deliver antigens to induce robust immune response. Based on these properties, exosomes could be used as a biological immunotherapy by enhancing adaptive immune response to inhibit hepatitis D virus replication. Our research may provide a new therapeutic strategy to eradicate HDV in the future.

HOXA10 knockdown inhibits proliferation, induces cell cycle arrest and apoptosis in hepatocellular carcinoma cells through HDAC1.

BACKGROUND: Homeobox A10 (HOXA10) has been implicated in the development and progression of various human cancers. However, the precise biological functions of HOXA10 in hepatocellular carcinoma (HCC) have not been defined. METHODS: In this study, we examined mRNA expression by quantitative real-time PCR (qRT-PCR) of HOXA10 as well as histone deacetylase (HDAC) and protein levels by Western blot of HOXA10, HDAC1, Cyclin D1, proliferating cell nuclear antigen (PCNA), Survivin and p53 acetylation in HCC tissues and cell lines. We also assessed cell proliferation using Cell Counting Kit-8 (CCK-8) and analyzed cell cycle by flow cytometry. Furthermore, tumor growth of HCC cells in vivo was monitored using the nude mouse xenograft model. Finally, HDAC1 promoter activity and binding in HCC cell lines were detected by luciferase reporter assay and chromatin immunoprecipitation (ChIP), respectively. RESULTS: We uncovered the elevated expression of HOXA10 in HCC tissues compared to adjacent normal liver tissues. RNA interference-mediated knockdown of HOXA10 inhibited HCC cell proliferation both in vitro and in vivo. HOXA10 knockdown also induced cell cycle arrest at G0/G1 phase and apoptosis, which were accompanied with the reduced expression of Cyclin D1, PCNA and Survivin. Notably, HOXA10 knockdown enhanced p53 acetylation (Lys382), which is crucial to the activation of p53. Likewise, HOXA10 knockdown suppressed the transcription of HDAC1, a potential deacetylase for p53. In line with these observations, HDAC1 downregulation abrogated the effects of HOXA10 overexpression on proliferation, cell cycle progression, apoptosis and p53 acetylation, indicating the role of HDAC1 in mediating HOXA10 functions. CONCLUSION: Our results demonstrate that HOXA10 knockdown inhibits proliferation, induces cell cycle arrest and apoptosis in HCC cells by regulating HDAC1 transcription.

Plumbagin inhibits proliferation and induces apoptosis of hepatocellular carcinoma by downregulating the expression of SIVA.

Purpose: Plumbagin is thought to be a bioactive phytochemical drug and exerts an antitumor effect on various cancers. However, few studies focus on the antitumor activity of plumbagin on liver cancer. This study first investigated the antitumor activity of plumbagin on liver cancer and further investigated the molecular mechanism of its antitumor activity against hepatocellular carcinoma, both in vitro and in vivo. Methods: The antiproliferative activity of plumbagin was evaluated through CCK-8, EdU, and colony forming test. The cell cycle and apoptosis were then analyzed by flow cytometer. Western blot was used to detect the expression of apoptosis related protein, SIVA, and mTOR pathway. RNA-seq was performed to determine the gene expression profiles and overexpressed or knocked down SIVA to validate its role in plumbagin's antitumor activity. Regarding animal experiment, a xenograft model in BALB/c nude mice was built using LM3-Luci cells. Then bioluminescence imaging and further immunohistochemistry were performed to study the antitumor activity and the expression of SIVA and mTOR in the plumbagin-treated group. Results: Plumbagin can inhibit proliferation and induce apoptosis of liver cancer cells in vitro. Further experiment demonstrated that plumbagin could inhibit the expression of SIVA and subsequently downregulate the mTOR signaling pathway, and upregulating the expression of SIVA will alleviate the antitumor activity of plumbagin on liver cancer, which confirmed the important role of the SIVA/mTOR signaling pathway in the antitumor activity of plumbagin. In vivo bioluminescence imaging showed a decreased signal in the plumbagin-treated group, and further immunohistochemistry demonstrated that plumbagin could inhibit the SIVA/mTOR signaling pathway in tumor tissues. Conclusion: Our promising results showed that plumbagin could inhibit proliferation and induce apoptosis of hepatic cancer through inhibiting the SIVA/mTOR signaling pathway for the first time, which indicated that plumbagin might be a good candidate against liver cancer.

Tripartite motif-containing protein 7 regulates hepatocellular carcinoma cell proliferation via the DUSP6/p38 pathway.

Tripartite motif-containing protein 7 (TRIM7), which is involved in the biosynthesis of glycogen, has been reported to drive lung tumorigenesis. In the present study, we aimed to examine the expression, roles and underlying molecular mechanisms of TRIM7 in hepatocellular carcinoma (HCC) development. Real-time PCR and immunohistochemical staining were performed to test the expression of TRIM7 in HCC tissues. Cell proliferation, cell cycle and tumorigenicity experiments were conducted to determine the function of TRIM7. The results showed that TRIM7 expression was elevated in human HCC tissues and that TRIM7 expression was significantly associated with tumor size, pTNM stage, serum α-fetoprotein (AFP) concentration, serum hepatitis B virus (HBV) DNA copy number and overall survival (OS) of HCC patients. TRIM7 knockdown inhibited the proliferation of HCC cells in vitro and in vivo. TRIM7 knockdown also induced a G1/S checkpoint in HCC cell lines. Additionally, TRIM7 knockdown led to decreased phosphorylated p38 (p-p38) and increased expression of p53 and p21. Ectopic expression of TRIM7 promoted HCC cell proliferation, cell cycle progression and p38 activation, but not in the presence of the p38 inhibitor SB203580. Moreover, TRIM7 overexpression enhanced the polyubiquitination and degradation of dual specificity phosphatase 6 (DUSP6). DUSP6 overexpression abolished the promotional effect of TRIM7 overexpression on HCC cell proliferation and the activation of p38. Furthermore, HBV X protein (HBx), a protein coded by HBV, was demonstrated to upregulate TRIM7 expression. Collectively, TRIM7 overexpression may contribute to the highly proliferative characteristics of HCC cells, and targeting TRIM7 might be a potential strategy for HCC treatment.

An engineered novel lentivector specifically transducing dendritic cells and eliciting robust HBV-specific CTL response by upregulating autophagy in T cells.

Dendritic cells (DCs) play a predominant role in initiating cell immune responses. Here we generated a DC-targeting lentiviral vector (LVDC-UbHBcAg-LIGHT) and evaluated its capacity to elicit HBV-specific cytotoxic T lymphocyte (CTL) responses. DC-SIGN-mediated specific transduction using this construct was confirmed in DC-SIGN-expressing 293T cells and ex vivo-cultured bone marrow cells. LVDC-UbHBcAg-LIGHT-loaded DCs were highly effective in inducing HBV-specific CTLs. Mechanistic studies demonstrated autophagy blocking led to a significant increase in apoptosis and obvious inhibition of CD8 + T cells entry into S-phase, correspondingly attenuated LVDC-UbHBcAg-LIGHT-loaded DC-induced T cell responses. This observation was supported by accumulation of pro-apoptotic proteins and the main negative cell cycle regulator-CDKN1B that otherwise would be degraded in activated T cells where autophagy preferentially occured. Our findings revealed an important role of autophagy in the activation of T cells and suggested LVDC-UbHBcAg-LIGHT may potentially be used as a therapeutic strategy to combat persistent HBV infection with higher security.

TRIM52 up-regulation in hepatocellular carcinoma cells promotes proliferation, migration and invasion through the ubiquitination of PPM1A.

BACKGROUND: Many tripartite motif (TRIM) family proteins have been reported to be of great importance in the initiation and progression in hepatocellular carcinoma (HCC). However, the biological role and regulatory mechanism of tripartite motif containing 52 (TRIM52) in HCC development and progression are poorly defined. METHODS: Immunohistochemistry (IHC), quantitative real-time PCR (qRT-PCR) or Western blot analysis was used to detect TRIM52, p21, matrix metalloproteinase 2 (MMP2), protein phosphatase, Mg2+/Mn2+ dependent 1A (PPM1A), p-Smad2/3 and Smad2/3 levels in HCC tissues and cell lines. HCC cell proliferation and cell cycle were measured by Cell Counting Kit-8 (CCK-8) and flow cytometry analysis, respectively. HCC cell migration and invasion were measured by Transwell assay. Tumor growth of HCC cells in vivo was measured using the nude mouse xenograft model. The correlation between TRIM52 and PPM1A was measured by co-immunoprecipitation (Co-IP) and ubiquitination analysis in vitro. RESULTS: TRIM52 was significantly up-regulated in the HCC tissues in comparison with the adjacent non-tumor hepatic tissues. TRIM52 was also up-regulated in HCC cell lines (MHCC-97H and MHCC-97L cells) compared with normal human liver cell line LO2. TRIM52 down-regulation by RNA interfering in MHCC-97H cells enhanced inhibition of cell proliferation, migration and invasion. TRIM52 down-regulation also induced MHCC-97H cells arrest in G0-G1 phase cell cycle and inhibited MHCC-97H cell growth in the nude mice. However, TRIM52 up-regulation in MHCC-97L cells promoted cell proliferation, migration and invasion. Furthermore, TRIM52 down-regulation significantly increased p21 and PPM1A expression, but inhibited MMP2 expression and induced Smad2/3 dephosphorylation in MHCC-97H cells, which were reversed by TRIM52 up-regulation in MHCC-97L cells. TRIM52 was found interacted with PPM1A and TRIM52 down-regulation inhibited the ubiquitination of PPM1A. Importantly, PPM1A up-regulation in MHCC-97L cells significantly suppressed TRIM52-mediated enhancement on cell proliferation, invasion and migration. CONCLUSIONS: Our findings suggest that TRIM52 up-regulation promotes proliferation, migration and invasion of HCC cells through the ubiquitination of PPM1A.

Delivery of Tapasin-modified CTL epitope peptide via cytoplasmic transduction peptide induces CTLs by JAK/STAT signaling pathway in vivo.

Hepatitis B virus (HBV)-specific cytotoxic T lymphocytes (CTLs) play a vital role in viral control and clearance. Recent studies have elucidated that Tapasin, an endoplasmic reticulum chaperone, is a well-known molecule that appears to be essential in peptide-loading process. The Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway plays an important role in immune response regulation and cytokines secretion. We have previously verified that fusion protein CTP-HBcAg18-27-Tapasin could facilitate the maturation of bone marrow derived dendritic cells and enhance specific CTLs responses in vitro, which might be associated with the activation of JAK/STAT signaling pathway. To further explore whether JAK/STAT signaling pathway participated in specific immune responses mediated by CTP-HBcAg18-27-Tapasin, we suppressed the JAK/STAT pathway with pharmacological inhibitor (AG490) in vivo. Our studies showed that the number of IFN-γ+-CD8+ T cells was decreased significantly compared with other groups after being blocked by AG490. The percentage of IFN-γ+-CD4+ T cells and IL-2-CD4+ T cells was also decreased. Moreover, lower expression levels of Jak2, Tyk2, STAT1, and STAT4 were detected in AG490 group. In addition, the secretion levels of Th1-like cytokines were decreased and a weaker specific T-cell response was observed in AG490 group. Furthermore, the levels of HBV DNA and HBsAg in serum and expression levels of HBsAg and HBcAg in liver tissues were elevated after this pathway was inhibited in HBV transgenic mice. These results demonstrate that the JAK/STAT signaling pathway participates in Th1-oriented immune response induced by CTP-HBcAg18-27-Tapasin and this might provide a theoretical basis for HBV immunotherapy.

Diverse Changes of Circulating Fibroblast Growth Factor 21 Levels in Hepatitis B Virus-Related Diseases.

Fibroblast growth factor 21 (FGF21), a stress-induced hormone in the liver, has been shown the protective functions in pathological conditions. The study investigated the association of circulating FGF21 with hepatitis B virus (HBV) infection and its related diseases. Serum FGF21 levels were measured in 33 acute hepatitis B (AHB), 75 chronic hepatitis B (CHB) and 66 CHB patients with advanced liver diseases including liver cirrhosis, acute-on-chronic liver failure (ALCF) and hepatocellular carcinoma (HCC) together with 200 age- and BMI-matched healthy controls. FGF21 levels were significantly increased in AHB patients and rapidly returned to normal levels after treatment. FGF21 levels reflected the degree of liver injury caused by AHB. However, serum FGF21 levels were decreased in CHB patients especially in those who developed cirrhosis and were associated with hepatic protein synthesis capacity. Serum FGF21 in CHB patients were increased with the occurrence of ACLF. Notably, in CHB patients who developed HCC, serum FGF21 exhibited a dramatic increase, which may provide important information on monitoring tumorigenesis in CHB patients. In conclusion, we revealed the diverse changes of circulating FGF21 in HBV-related diseases. FGF21 may be a useful biomarker in monitoring the tumorigenesis in patients with CHB.

Tripartite Motif Containing 52 (TRIM52) Promotes Cell Proliferation in Hepatitis B Virus-Associated Hepatocellular Carcinoma.

BACKGROUND Chronic hepatitis B virus (HBV) infection is the major cause of hepatocellular carcinoma (HCC). HBV X protein (HBx) plays a crucial role in the development of HCC. Moreover, many tripartite motif (TRIM) family proteins exert diverse biological functions in hepatocarcinogenesis. However, as a novel member of this family, the specific effect of TRIM52 is still largely obscure. In the present study, we investigated the expression and function of TRIM52 in HBV-associated HCC. MATERIAL AND METHODS Fluorescence quantitative polymerase chain reaction (FQ-PCR) was performed to detect the HBV DNA levels in the peripheral blood of HCC patients. Quantitative real-time PCR (qRT-PCR) and Western blot analysis were performed to detect the expression of TRIM52, HBx, and NF-κB p65. HBx-pcDNA3.1 and TRIM52-shRNA were used to induce HBx ectopic expression and TRIM52 silencing, respectively. Pyrrolidine dithiocarbamate (PDTC) was used to block the activation of NF-κB. Cell proliferation was detected using the Cell Counting Kit-8 (CCK-8) assay. RESULTS TRIM52 expression was up-regulated together with HBx in HBV-associated HCC tissues. Ectopic expression of HBx elevated TRIM52 expression in HepG2 cells. TRIM52 silencing repressed the proliferation of HepG2.2.15 cells. Moreover, NF-κB p65 expression was increased in HCC cell lines. Blocking NF-κB activation with PDTC suppressed TRIM52 expression and attenuated the viability of HepG2.2.15 cells. CONCLUSIONS These findings indicate that TRIM52 can promote cell proliferation and HBx may regulate TRIM52 expression via the NF-κB signaling pathway in HBV-associated HCC.

Correlation between low tapasin expression and impaired CD8+ T­cell function in patients with chronic hepatitis B.

Recent studies have demonstrated that chronic hepatitis B virus (HBV) infection is associated with reduced antigen­presenting capacity and insufficient cytotoxic T lymphocyte (CTL) production. The molecular chaperone tapasin mediates binding of the transporter associated with antigen processing (TAP), and has an important role in endogenous antigen processing and presentation, and the induction of specific CTL responses. The present study aimed to determine whether tapasin is associated with chronic HBV (CHB) infection. The mRNA expression levels of tapasin were detected in peripheral blood mononuclear cells from 27 patients with CHB, 20 patients with acute HBV (AHB) and 26 healthy controls by reverse transcription­quantitative polymerase chain reaction. In addition, CD8+ T immune responses were evaluated in all groups, and the correlation between tapasin expression and CD8+ responses was analyzed. The results demonstrated that the mRNA expression levels of tapasin were significantly downregulated in patients with CHB compared with in healthy controls and patients with AHB. Furthermore, the apoptotic rate of CD8+ T cells was increased in patients with CHB compared with in the other two groups. The percentage of interferon (IFN)­Î³+CD8+ T cells was reduced in patients with CHB compared with in patients with AHB and healthy controls, and serum cytokine levels (IFN­Î³, interleukin­2 and tumor necrosis factor­α) were generally low in patients with CHB. Furthermore, the mRNA expression levels of tapasin were positively correlated with IFN­Î³ production by CD8+ T cells, and were inversely correlated with the apoptotic ratio of CD8+ T cells. These results indicate that decreased expression of tapasin may be closely associated with CHB, and suggest an important role for tapasin in the pathogenesis of CHB.

Lentiviral vector encoding ubiquitinated hepatitis B core antigen induces potent cellular immune responses and therapeutic immunity in HBV transgenic mice.

Predominant T helper cell type 1 (Th1) immune responses accompanied by boosted HBV-specific cytotoxic T lymphocyte (CTL) activity are essential for the clearance of hepatitis B virus (HBV) in chronic hepatitis B (CHB) patients. Ubiquitin (Ub) serves as a signal for the target protein to be recognized and degraded through the ubiquitin-proteasome system (UPS). Ubiquitinated hepatitis B core antigen (Ub-HBcAg) has been proved to be efficiently degraded into the peptides, which can be presented by major histocompatibility complex (MHC) class I resulting in stimulating cell-mediated responses. In the present study, lentiviral vectors encoding Ub-HBcAg (LV-Ub-HBcAg) were designed and constructed as a therapeutic vaccine for immunotherapy. HBcAg-specific cellular immune responses and anti-viral effects induced by LV-Ub-HBcAg were evaluated in HBV transgenic mice. We demonstrated that immunization with LV-Ub-HBcAg promoted the secretion of cytokines interleukin-2 (IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α), generated remarkably high percentages of IFN-γ-secreting CD8(+) T cells and CD4(+) T cells, and enhanced HBcAg-specific CTL activity in HBV transgenic mice. More importantly, vaccination with LV-Ub-HBcAg could efficiently decreased the levels of serum hepatitis B surface antigen (HBsAg), HBV DNA and the expression of HBsAg and HBcAg in liver tissues of HBV transgenic mice. In addition, LV-Ub-HBcAg could upregulate the expression of T cell-specific T-box transcription factor (T-bet) and downregulate the expression of GATA-binding protein 3 (GATA-3) in spleen T lymphocytes. The therapeutic vaccine LV-Ub-HBcAg could break immune tolerance, and induce potent HBcAg specific cellular immune responses and therapeutic effects in HBV transgenic mice.

Tuberculosis ineection might increase the risk f invasive candidiasis in an immmunocompetent patient / Tuberculose pode aumentar o risco de candidíase invasiva em paciente imunocompetente

Deep Candida infections commonly occur in immunosuppressed patients. A rare case of a multiple deep organ infection with Candida albicans and spinal tuberculosis was reported in a healthy young man. The 19-year-old man complained of month-long fever and lower back pain. He also had a history of scalded mouth syndrome. Coinfection with Mycobacterium tuberculosis and Candida albicans was diagnosed using the culture of aspirates from different regions. Symptoms improved considerably after antifungal and antituberculous therapy. This case illustrates that infection with tuberculosis might impair the host's immune system and increase the risk of invasive candidiasis in an immunocompetent patient.

As infecções profundas por Candida ocorrem geralmente em pacientes imunossuprimidos. Relatamos caso raro de infecções profundas em múltiplos órgãos por Candida albicans e neuro tuberculose em homem jovem saudável. Um jovem de 19 anos de idade queixou-se de febre e lombalgia há um mês. Relatava ainda histórico de síndrome da boca escaldada. Foi diagnosticada co-infecção por Mycobacterium tuberculosis e Candida albicans em cultura do aspirado de diferentes regiões do organismo. Os sintomas melhoraram significativamente após a terapia antifúngica e antituberculosa. Este caso é apresentado para mostrar que a tuberculose pode prejudicar o sistema imune do hospedeiro e aumentar o risco de candidíase invasiva em paciente imunocompetente.

Vaccination with ubiquitin-hepatitis B core antigen-cytoplasmic transduction peptide enhances the hepatitis B virus-specific cytotoxic T-lymphocyte immune response and inhibits hepatitis B virus replication in transgenic mice.

Chronic hepatitis B virus (HBV) infection is characterized by functionally impaired type 1 T-helper cell (Thl) immunity and poor HBV­specific T­cell responses. Ubiquitin (Ub), a highly conserved small regulatory protein, commonly serves as a signal for target proteins that are recognized and degraded in proteasomes. The rapid degradation of Ub­mediated antigens results in efficient stimulation of cell­mediated immune responses. Thus, the Ub­HBV core antigen (HBcAg)­cytoplasmic transduction peptide (CTP) fusion protein was designed for specific delivery of a foreign modified antigen to the cytoplasm of antigen­presenting cells. HBV transgenic mice were used to determine whether Ub­HBcAg­CTP would restore HBV­specific immune responses and anti­viral immunity in these animals. The results demonstrated that synthesized Ub­HBcAg­CTP not only significantly increased the levels of interleukin­2 and interferon (IFN)­Î³ compared with those in the HBcAg­CTP, IFN­α, Ub­HBcAg, HBcAg and phosphate­buffered saline groups, but additionally induced the highest IFN­Î³+ CD8+ T­cell numbers and HBV­specific cytotoxic T lymphocyte (CTL) responses, indicating a strong immune response. In addition, enhancement of specific CTL activity provoked by the fusion protein reduced hepatitis B surface antigen (HBsAg) and HBV DNA serum levels and diminished the expression of HBsAg and HBcAg in liver tissue of HBV transgenic mice, suggesting that there was a therapeutic effect. In conclusion, the present study provided evidence that Ub­HBcAg­CTP activated the Th1­dependent immunity, triggered functional T cell responses and subsequently inhibited viral replication in HBV transgenic mice. These observations suggested that the fusion protein may represent an innovative and promising candidate for active immunotherapy during chronic and persistent HBV.

Ubiquitin-modified hepatitis B virus core antigen effectively facilitates antigen presentation and enhances cytotoxic T lymphocyte activity via the cytoplasmic transduction peptide in vitro.

Cluster of differentiation (CD)8+ cytotoxic T lymphocytes (CTLs) have a key role in the elimination of hepatitis B virus (HBV)-infected cells. Ubiquitin (Ub) functions as a marker for protein degradation, which may promote the generation of peptides appropriate for major histocompatibility complex class I presentation, while the HBV core antigen (HBcAg) possesses marked immunogenic properties. However, it remains to be elucidated whether Ub-modified HBcAg is able to effectively elicit significant CD8+ CTL activity. In order to address this issue, a prokaryotic vector was constructed to express the Ub-HBcAg-cytoplasmic transduction peptide (CTP). The fusion protein was successfully expressed and subsequently pulsed into bone-marrow-derived dendritic cells (DCs). It was confirmed that with assistance from the cell­penetrating properties of CTP, the fusion protein was able to directly penetrate into the cytoplasm of DCs. The results revealed that the Ub-HBcAg-CTP fusion protein not only increased the expression of surface molecules in DCs and cytokine secretion from proliferating T cells, but also induced T cells to differentiate into specific CTLs and enhanced their antiviral ability. In conclusion, the Ub-HBcAg-CTP fusion protein promoted DC maturation, enhanced the presentation of targeting antigens and efficiently induced HBcAg­specific CTL immune responses in vitro.

Hepatitis B virus-associated glomerulonephritis in HBsAg serological-negative patients.

OBJECTIVE: The main objective of this study is to report the clinical and pathological characteristics of hepatitis B virus (HBV)-associated glomerulonephritis (HBV-GN) in serum hepatitis B surface antigen (HBsAg)-negative patients in China. HBV-GN is caused by the HBV's attack on the kidney tissues, but definitive diagnostic criteria are still lacking. The diagnostic criteria used in China require HBsAg positivity in the serum, but research on occult HBV infection has shown that HBV infection is also found in serum HBsAg-negative patients. Clinical and pathological characterization of HBV-GN in serum HBsAg-negative patients is required. MATERIALS AND METHODS: Serologic and clinical findings and pathological characteristics of renal tissues in 18 HBV-GN patients (11 men and seven women) with serum HBsAg negativity were analyzed retrospectively. Thirty-three HBV-GN patients with serum HBsAg positivity and 59 patients with membranous nephropathy (MN) without any HBV infection markers in serum and renal tissues during the same period were included as controls. RESULTS: Among the 18 HBsAg-negative patients with GN, 12 had HBsAb positivity in their sera. None of the patients was positive for serum HBeAg. Proteinuria was the major clinical manifestation and the renal histopathology was characterized as MN. Immune fluorescence deposits in renal tissues consisted mainly of HBsAg. The degree of renal injury and the decrease in the C3 level were less than those in HBsAg-positive patients and idiopathic membranous nephropathy patients. CONCLUSION: We propose to use the HBV marker in renal tissues as a new diagnostic criterion for HBV-GN. If MN patients have HBV marker positivity in renal tissues, HBV-GN can be diagnosed even without HBsAg in the serum. This would improve the diagnostic accuracy and potential treatment efficiency.