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In vitro use of autologous dendritic cells improves detection of T cell responses to hepatitis B virus (HBV) antigens.
Dez. 2008 | Carotenuto, Patrizia; Artsen, Andrè; Niesters, Hubert G; Osterhaus, Albert D; Pontesilli, Oscar
T lymphocyte responses to hepatitis B virus (HBV) core antigen (HBcAg) are vigorous and easily detectable in vitro during recovery from acute hepatitis B but significantly weaker in patients with chronic HBV infection. In contrast, T cell responses to hepatitis B surface antigen (HBsAg) are almost undetectable during infection and even in a substantial fraction of subjects receiving vaccination with HBsAg. The aim of this study was to investigate whether the use of dendritic cells (DCs) in an in vitro assay could increase the detection of HBV-specific T cells in these conditions. Autologous monocyte-derived DCs, compared to direct HBsAg addition to the cultures, increased the stimulation of HBs- specific T cells. These were detected in 73% of healthy subjects who had recently received hepatitis B vaccine and in 43% of patients recovering from acute hepatitis B. Likewise, proliferation in response to DC-presented HBcAg was detected in both CD4(+) and CD8(+) T cells from the majority of chronic hepatitis B patients. A longitudinal evaluation of HBc-specific T cell responses during and after a 1-year treatment with pegylated interferon (IFN)-alpha showed that HBc-specific CD4(+) T cell responses had no correlation with sustained virus suppression whereas CD8(+) T cell responses were more frequently detected in patients able to control HBV replication after therapy interruption. The use of autologous DCs as antigen-presenting cells appears applicable to clinically relevant in vitro evaluation of T cell responses, particularly in those conditions characterized by low frequency of circulating antigen-specific cells and suboptimal in vivo activation. PMID 19107973

Monocyte-derived dendritic cells from HCV-infected patients transduced with an adenovirus expressing NS3 are functional when stimulated with the TLR3 ligand poly(I:C).
Nov. 2008 | Echeverría, I; Zabaleta, A; Silva, L; Díaz-Valdés, N; Riezu-Boj, J I; Lasarte, J J; Borrás-Cuesta, F; Civeira, M P; Prieto, J; Sarobe, P
Dendritic cells (DC) transfected with an adenovirus encoding hepatitis C virus (HCV) NS3 protein (AdNS3) induce potent antiviral immune responses when used to immunize mice. However, in HCV infected patients, controversial results have been reported regarding the functional properties of monocyte-derived DC (MoDC), a cell population commonly used in DC vaccination protocols. Thus, with the aim of future vaccination studies we decided to characterize MoDC from HCV patients transfected with AdNS3 and stimulated with the TLR3 ligand poly(I:C). Phenotypic and functional properties of these cells were compared with those from MoDC obtained from uninfected individuals. PCR analysis showed that HCV RNA was negative in MoDC from patients after the culture period. Also, phenotypic analysis of these cells showed lower expression of CD80, CD86, and CD40, but similar expression of HLA-DR molecules as compared to MoDC from uninfected individuals. Functional assays of MoDC obtained from patients and controls showed a similar ability to activate allogeneic lymphocytes or to produce IL-12 and IL-10, although lower IFN-alpha levels were produced by cells from HCV patients after poly(I:C) stimulation. Moreover, both groups of MoDC induced similar profiles of IFN-gamma and IL-5 after stimulation of allogeneic T-cells. Finally, migration assays did not reveal any difference in their ability to respond to CCL21 chemokine. In conclusion, MoDC from HCV patients are functional after transduction with AdNS3 and stimulation with poly(I:C). These findings suggest that these cells may be useful for therapeutic vaccination in chronic HCV infection. PMID 18637068

Monocyte derived dendritic cells retain their functional capacity in patients following infection with hepatitis C virus.
Jan. 2008 | Barnes, E; Salio, M; Cerundolo, V; Francesco, L; Pardoll, D; Klenerman, P; Cox, A
Studies assessing the function of monocyte derived dendritic cells (MD-DC) in individuals with hepatitis C virus (HCV) infection have shown conflicting results. Impaired MD-DC function in chronic HCV infection would have important implications both for understanding the pathogenesis of HCV infection and in the use of autologous MD-DC in vaccination strategies. We determined the allostimulatory capacity of MD-DC in the same patient before and after HCV infection. Next, the phenotype, cytokine production and allostimulatory function of immature and mature MD-DC in individuals with persistent HCV infection were compared directly with MD-DC from healthy individuals. Finally, we assessed the ability of MD-DC to prime autologous naïve peptide specific CD8+ T cells using HLA-A2 class-I tetramers. DCs retained the same allostimulatory capacity before and following the establishment of persistent HCV infection. The surface phenotype and the amount of interleukin (IL)-10 and IL-12(p70) produced during DC maturation did not differ between HCV-infected individuals and healthy controls. Mature DCs from HCV-infected individuals performed comparably in an allogeneic MLR compared with healthy individuals. Mature MD-DC from HCV-infected individuals stimulated the expansion of peptide specific naïve CD8+ T cells. MD-DC from HCV-infected and healthy individuals are phenotypically indistinguishable and perform comparably in functional assays. PMID 18194173

Vaccination against hepatitis C virus with dendritic cells transduced with an adenovirus encoding NS3 protein.
Dez. 2007 | Zabaleta, Aintzane; Llopiz, Diana; Arribillaga, Laura; Silva, Leyre; Riezu-Boj, José Ignacio; Lasarte, Juan José; Borrás-Cuesta, Francisco; Prieto, Jesús; Sarobe, Pablo
Chronic infection by hepatitis C virus (HCV) is characterized by the absence of efficient antiviral T-cell responses. Thus, vaccination strategies to induce strong anti-HCV T-cell responses are of paramount importance for prophylactic and therapeutic purposes. Dendritic cells (DCs) are the most potent antigen presenting cells; therefore, immunization with these cells loaded with viral antigens offers a new approach for induction of antiviral immunity. Here we show that immunization with DCs transfected with an adenovirus encoding non-structural 3 protein, from HCV (AdNS3), induced multiepitopic CD4 T helper cell 1 (Th1) and CD8 T-cell responses in different mouse strains. These responses prevented the growth of a tumorexpressing HCV proteins, in short- and long-term experiments. Moreover, immunization with AdNS3-transfected DCs did not induce anti-adenoviral antibodies, as compared to direct immunization with AdNS3, but elicited T-cell responses even in the presence of pre-existing anti-adenoviral antibodies. Finally, responses induced by this protocol down-regulated the expression of HCV RNA in the liver. In conclusion, DCs transfected with AdNS3 may prove to be an efficient anti-HCV vaccine. PMID 17923840

Monocyte-derived dendritic cells from chronic HCV patients are not infected but show an immature phenotype and aberrant cytokine profile.
Aug. 2007 | Gelderblom, Huub C; Nijhuis, Laurens E J; de Jong, Esther C; te Velde, Anje A; Pajkrt, Dasja; Reesink, Henk W; Beld, Marcel G H M; van Deventer, Sander J H; Jansen, Peter L M
Chronic hepatitis C virus (HCV) infection is characterized by an insufficient immune response, possibly owing to impaired function of antigen-presenting cells such as myeloid dendritic cells (DCs). Therapeutic vaccination with in vitro generated DCs may enhance the immune response. Subsets of DCs can originate from monocytes, but the presence of HCV in monocytes that develop into DCs in vitro may impair DC function. Therefore, we studied the presence of HCV RNA in monocytes and monocyte-derived DCs from chronic HCV patients. PMID 17696933

Monocyte-derived dendritic cell function in chronic hepatitis C is impaired at physiological numbers of dendritic cells.
Mai 2007 | MacDonald, A J; Semper, A E; Libri, N A; Rosenberg, W M C
Monocyte-derived dendritic cells (MoDCs) are a promising cellular adjuvant for effector immune responses against tumours and chronic viral infections, including hepatitis C virus (HCV). If autologous DC therapeutic approaches are to be applied in persistent HCV infections in patients, it is important to have an unambiguous understanding of the functional status of the cell type used, namely MoDCs from patients with chronic hepatitis C (CHC) infection. Because of conflicting published reports of either impaired or normal MoDC function in CHC infection, we re-examined the ability of MoDCs from CHC and normal healthy donors (NHD) to mature to an inflammatory stimulus [tumour necrosis factor (TNF)-alpha] and their subsequent functional capabilities. Expression of maturation-associated phenotypic markers [human leucocyte antigen (HLA)-DR, CD83, CD86, CD40], allostimulatory capacity in mixed lymphocyte reactions (MLRs) and CD40-ligand-induced cytokine and chemokine generation were compared in CHC- versus NHD-MoDCs. TNF-alpha-stimulated CHC-MoDCs up-regulated phenotypic markers, but to significantly lower levels than NHD-MoDCs. At physiological ratios of DCs to T cells, CHC-MoDCs were less allostimulatory than NHD-MoDCs, but not when DC numbers were substantially increased. CHC- and NHD-MoDCs generated equivalent amounts of cytokines [TNF-alpha, interleukin (IL)-1beta, IL-6, IL-12p70, IL-15, IL-10] and chemokines [interferon-inducible protein (IP)-10, macrophage inflammatory protein (MIP)-1alpha, regulated upon activation, normal T expressed and secreted (RANTES)] after CD40 ligation. Because the functional defect was not apparent at high MoDC : T cell ratios, autologous MoDC therapy with sufficiently high numbers of DCs could, in theory, overcome any impairment of MoDC function in CHC. PMID 17362265

Hepatitis B virus (HBV) antigen-pulsed monocyte-derived dendritic cells from HBV-associated hepatocellular carcinoma patients significantly enhance specific T cell responses in vitro.
Jan. 2007 | Shi, M; Qian, S; Chen, W-W; Zhang, H; Zhang, B; Tang, Z-R; Zhang, Z; Wang, F-S
To investigate whether hepatitis B virus (HBV) antigen-pulsed monocyte-derived dendritic cells (MoDC) could mount a T cell response in hepatocellular carcinoma (HCC) patients associated with chronic HBV infection, peripheral blood mononuclear cells (PBMCs) from 36 HBV-associated HCC patients were induced into MoDC and pulsed with hepatitis B core antigen (HBcAg) and hepatitis B surface antigen (HBsAg), alone and in combination. Co-stimulatory molecules CD80, CD86 and CD40, as well as human leucocyte antigens D-related (HLA-DR) were found to express at the highest level on MoDC pulsed with HBcAg or HBsAg + HBcAg, at a median level on MoDC pulsed with HBcAg or HBsAg alone, and at the lowest level on non-antigen-pulsed MoDC. Interleukin (IL)-10 and IL-12 cytokines were released by antigen-pulsed MoDC at increased levels in the order: no-antigen < HBsAg < HBcAg < HBcAg + HBsAg. MoDC pulsed with HBcAg or HBsAg + HBcAg also had the strongest ability to stimulate autologous T cell proliferation and intracellular interferon (IFN)-gamma production. HBcAg- or HBsAg + HBcAg-pulsed MoDC could also induce HBV core peptide-specific CD8(+) T cell proliferation determined by tetramer staining. In addition, the antigen-pulsed MoDC were found to have a stronger capacity to produce IL-12 and induce T cell response in vitro for patients with higher alanine transaminase (ALT) levels than those with lower ALT levels, indicating that antigen pulse could substantially reverse the impaired function of MoDC in primary HCC patients with active chronic hepatitis B. In conclusion, HBV antigen-pulsed MoDC from HCC patients with chronic hepatitis B could induce HBV-specific T cell response in vitro. PMID 17223969

Induction of primary human T cell responses against hepatitis C virus-derived antigens NS3 or core by autologous dendritic cells expressing hepatitis C virus antigens: potential for vaccine and immunotherapy.
Mai 2006 | Li, Wen; Krishnadas, Deepa K; Li, Jie; Tyrrell, D Lorne J; Agrawal, Babita
Hepatitis C virus (HCV)-specific T cell responses have been suggested to play significant role in viral clearance. Dendritic cells (DCs) are professional APCs that play a major role in priming, initiating, and sustaining strong T cell responses against pathogen-derived Ags. DCs also have inherent capabilities of priming naive T cells against given Ags. Recombinant adenoviral vectors containing HCV-derived Core and NS3 genes were used to endogenously express HCV Core and NS3 proteins in human DCs. These HCV Ags expressing DCs were used to prime and stimulate autologous T cells obtained from uninfected healthy donors. The DCs expressing HCV Core or NS3 Ags were able to stimulate T cells to produce various cytokines and proliferate in HCV Ag-dependent manner. Evidence of both CD4(+) and CD8(+) T cell responses against HCV Core and NS3 generated in vitro were obtained by flow cytometry and Ab blocking experiments. Further, in secondary assays, the T cells primed in vitro exhibited HCV Ag-specific proliferative responses against recombinant protein Ags and also against immunodominant permissive peptide epitopes from HCV Ags. In summary, we demonstrate that the dendritic cells expressing HCV Ags are able to prime the Ag-specific T cells from uninfected healthy individuals in vitro. These studies have implications in designing cellular vaccines, T cell adoptive transfer therapy or vaccine candidates for HCV infection in both prophylactic and therapeutic settings. PMID 16670315

Vaccination with protein-transduced dendritic cells elicits a sustained response to hepatitis C viral antigens.
Feb. 2006 | Kuzushita, Noriyoshi; Gregory, Stephen H; Monti, Nola A; Carlson, Rolf; Gehring, Stephan; Wands, Jack R
Professional antigen-presenting dendritic cells are capable of eliciting a vigorous antiviral response in naive T cells. The administration of antigen-loaded dendritic cells offers a potential approach to induce high-level immunity against hepatitis C virus. PMID 16472599

Novel vaccines for the treatment of chronic HBV infection based on mycobacterial heat shock protein 70.
Feb. 2006 | Peng, Mingli; Chen, Min; Ling, Ning; Xu, Hongmei; Qing, Yuling; Ren, Hong
Immunogenic peptide-based vaccines can raise significant cellular immune responses. Although cytotoxic T lymphocytes (CTL) peptide epitopes are generally poor immunogens, heat shock protein 70 from Mycobacterium tuberculosis (TBhsp70) can overcome this problem since it is a potent adjuvant that links innate and adaptive immune responses. Our goal is to use TBhsp70 as an adjuvant for development of therapeutic vaccines for chronic Hepatitis B virus infection (HBV). To this end, we genetically fused the HBV core 18-27 peptide (HBcAg((18-27))) as a CTL epitope to the C-terminus of TBhsp70 and expressed the resulting protein in methylotropic yeast Pichia pastoris GS115. At the same time, the TBhsp70-HBcAg((18-27)) peptide complex was reconstituted in vitro. We investigated whether TBhsp70-peptide complex and TBhsp70-peptide fusion protein could generate antigen specific CTL responses in vitro. Dendritic cells (DC) from HLA-A2(+) chronic HBV infection and healthy control pulsed with two vaccines were studied phenotypically by FACS analyses and functionally by cytokine release, and HBV-specific CTL response. Our results demonstrate that two vaccines can activate DC of chronic HBV infection and healthy control by upregulation CD40 and CD86, high production of IL-12p70 and TNF-alpha. Furthermore, autologous T cells with DC stimulated by two vaccines can produce IFN-gamma and generate HBV-specific CTL response. However, capacity for CTL response and cytokines production from HBV infections remained inferior to that of healthy controls. Thus, the strategy of utilizing TBhsp70 may provide a novel design for the development of prophylactic and therapeutic vaccines. PMID 16446013

Dendritic cells pulsed with hepatitis C virus NS3 protein induce immune responses and protection from infection with recombinant vaccinia virus expressing NS3.
Dez. 2005 | Yu, Hong; Huang, Hui; Xiang, Jim; Babiuk, Lorne A; van Drunen Littel-van den Hurk, Sylvia
Infections with Hepatitis C virus (HCV) pose a serious health problem worldwide. In this study, the hypothesis that adoptive transfer of dendritic cells (DCs) pulsed with HCV NS3 protein and matured with an oligodeoxynucleotide (ODN) containing CpG motifs (CpG) ex vivo would initiate potent HCV-specific protective immune responses in vivo was tested. NS3 protein was efficiently transduced into DCs and treatment of DCs with CpG ODN induced phenotypic maturation and specifically increased the expression of CD40. DCs matured with CpG ODN produced higher interleukin 12 levels and a stronger allogeneic T-cell response compared with untreated DCs. Notably, there were no differences between NS3-pulsed DCs and DCs pulsed with a control protein with respect to phenotype, cytokine production or mixed lymphocyte reaction, indicating that transduction with NS3 protein did not impair DC functions. Compared with the untreated NS3-pulsed DCs, the NS3-pulsed DCs matured with CpG ODN induced stronger cellular immune responses including enhanced cytotoxicity, higher interferon-gamma production and stronger lymphocyte proliferation. Upon challenge with a recombinant vaccinia virus expressing NS3, all mice immunized with NS3-pulsed DCs showed a significant reduction in vaccinia virus titres when compared with mock-immunized mice. However, the NS3-pulsed DCs matured with CpG ODN induced higher levels of protection compared with the untreated NS3-pulsed DCs. These data are the first to show that NS3-pulsed DCs induce specific immune responses and provide protection from viral challenge, and also demonstrate that CpG ODNs, which have a proven safety profile, would be useful in the development of DC vaccines. PMID 16361412

Natural killer cells in hepatitis C virus infection: from innate immunity to adaptive immunity.
Okt. 2005 | Takehara, Tetsuo; Hayashi, Norio
Natural killer (NK) cells are specialized lymphocytes that provide a first line of defense through their ability to kill pathogen-infected cells and transformed cells. The function of NK cells is regulated by a fine balance of inhibitory and activating signals, which are mediated by a diverse array of cell-surface receptors. We recently found that expression of the inhibitory receptor CD94/NKG2A is up-regulated on NK cells in patients with chronic hepatitis C. HLA-E, a ligand for NKG2A, was expressed in all human hepatoma cell lines tested as well as in nontransformed hepatocytes, but not in K562 cells, a classic NK-sensitive target. NK cells isolated from patients with chronic hepatitis C (HCV-NK) were less capable of killing hepatoma cells and of producing interferon-gamma in response to hepatoma cells than those from healthy donors, whereas there was no significant difference in NK responsiveness toward K562 cells. Of note is the finding that maturation and activation of monocyte-derived dendritic cells were negatively modulated in the presence of HCV-NK and hepatoma cells, which were restored by the addition of anti-NKG2A antibody during the coculture of HCV-NK and hepatoma cells. Research revealed that dendritic cells recognize danger signals from microorganisms by monitoring pathogen-associated molecular patterns via Toll-like receptors. Our findings have shed light on NK receptors as an important interface that transmits danger signals from abnormal cells to immune systems. Aberrant expression of CD94/NKG2A should have negative impact on innate resistance and subsequent adaptive immunity toward HCV-infected or transformed cells in chronic hepatitis C. PMID 16234066

Prophylactic and therapeutic vaccination with dendritic cells against hepatitis C virus infection.
Okt. 2005 | Encke, J; Findeklee, J; Geib, J; Pfaff, E; Stremmel, W
Antigen uptake and presentation capacities enable DC to prime and activate T cells. Recently, several studies demonstrated a diminished DC function in hepatitis C virus (HCV) infected patients showing impaired abilities to stimulate allogenic T cells and to produce IFN-gamma in HCV infected patients. Moreover, DC of patients who have resolved HCV infection behave like DC from healthy donors responding to maturation stimuli, decrease antigen uptake, up-regulate expression of appropriate surface marker, and are potent stimulators of allogenic T cells. A number of studies have demonstrated in tumour models and models of infectious diseases strong induction of immune responses after DC vaccination. Because DC are essential for T-cell activation and since viral clearance in HCV infected patients is associated with a vigorous T-cell response, we propose a new type of HCV vaccine based on ex vivo stimulated and matured DC loaded with HCV specific antigens. This vaccine circumvents the impaired DC maturation and the down regulated DC function of HCV infected patients in vivo by giving the necessary maturation stimuli and the HCV antigens in a different setting and location ex vivo. Strong humoral and cellular immune responses were detected after HCV core DC vaccination. Furthermore, DC vaccination shows partial protection in a therapeutic and prophylactic model of HCV infection. In conclusion, mice immunized with HCV core pulsed DC generated a specific antiviral response in a mouse HCV challenge model. Our results indicate that HCV core pulsed DC may serve as a new modality for immunotherapy of HCV especially in chronically infected patients. PMID 16232225

Uptake and presentation of hepatitis C virus-like particles by human dendritic cells.
Apr. 2005 | Barth, Heidi; Ulsenheimer, Axel; Pape, Gerd R; Diepolder, Helmut M; Hoffmann, Marco; Neumann-Haefelin, Christoph; Thimme, Robert; Henneke, Philipp; Klein, Reinhild; Paranhos-Baccalà, Glaucia; Depla, Erik; Liang, T Jake; Blum, Hubert E; Baumert, Thomas F
Hepatitis C virus (HCV) is a major cause of chronic hepatitis worldwide. Interaction of dendritic cells (DCs) with viral particles may play an important role in the immunopathogenesis of HCV infection. Since the synthesis or purification of infectious virions is limited, we used HCV-like particles (HCV-LPs) to study the interaction of HCV with human DCs. Immature DCs exhibited an envelope-specific and saturable binding of HCV-LPs, indicating receptor-mediated DC-HCV-LP interaction. Confocal microscopy revealed that HCV-LPs were rapidly taken up by DCs in a temperature-dependent manner. Competition experiments demonstrated that C-type lectins such as mannose receptor or DC-SIGN (DC-specific intercellular adhesion molecule 3-grabbing nonintegrin) were not sufficient for mediating HCV-LP binding. HCV-LP uptake was followed by DC activation. DCs pulsed with HCV-LPs stimulated HCV core-specific CD4(+) T cells, indicating that uptake of HCV-LPs by DCs leads to antigen processing and presentation on major histocompatibility complex (MHC) class II molecules. Finally, HCV-LP-derived antigens were efficiently cross-presented to HCV core-specific CD8(+) T cells. These findings demonstrate that HCV-LPs represent a novel model system to study HCV-DC interaction allowing definition of the molecular mechanisms of HCV uptake, DC activation, and antigen presentation to T cells. Furthermore, HCV-LP-mediated DC activation and efficient antigen presentation may explain the marked immunogenicity of HCV-LPs in vivo. PMID 15657184

Production and efficacy of a dendritic cell-based therapeutic vaccine for murine chronic hepatitis B virus carrierer.
Juli 2004 | Akbar, S M F; Furukawa, Shinya; Hasebe, Aki; Horiike, Norio; Michitaka, Kojiro; Onji, Morikazu
Recently a new field of immunological research and clinical application of vaccines for therapeutic purposes (vaccine therapy) has been developed for treating several chronic viral infections including chronic hepatitis B virus (HBV) infection. Administration of vaccine containing hepatitis B surface antigen (HBsAg) for 1 year has resulted in negative HBsAg and development of antibody to HBsAg (anti-HBs) in some, but not in all, HBV transgenic mouse (HBV-Tg). In order to develop more potent regimen of vaccine therapy for chronic HBV carrier, we prepared a dendritic cell (DC)-based therapeutic vaccine and evaluated their therapeutic potential in HBV-Tg. DCs were isolated from single cell suspensions of murine spleen cells by collagenase digestion, density centrifugation and depletion of lymphocytes. Spleen DCs were cultured with HBsAg (100 microg) for 24 h to produce HBsAg-pulsed DCs. HBV-Tg expressing HBsAg and HBV DNA in the sera were randomly assigned to receive either HBsAg-pulsed DCs (n = 20) or unpulsed DC (n = 20) or vaccine containing HBsAg (n = 39) or complete Freund's adjuvant (n = 20) or left untreated (n = 20). Only two intraperitoneal injections of HBsAg-pulsed DCs resulted in negative HBsAg and production of anti-HBs in the sera in all HBV-Tg (n = 20). However, administration of un-pulsed DCs (n = 20) or vaccine containing HBsAg (n = 39) or only complete Freund's adjuvant did not induce negative HBsAg or production of anti-HBs in any HBV-Tg within 6 months of therapy commencement. Taken together, this study showed that HBsAg-pulsed DCs represent a highly potent therapeutic vaccine for chronic HBV infection and inspire optimism of using this vaccine in clinical conditions. A clinical trial of HBsAg-pulsed DC in patients with chronic hepatitis B is warranted. PMID 15254781

Prospects for dendritic cell vaccination in persistent infection with hepatitis C virus.
Mai 2004 | Gowans, Eric J; Jones, Kathryn L; Bharadwaj, Mandvi; Jackson, David C
Although hepatitis C virus (HCV) is classified in the Hepacivirus genus in the family Flaviviridae, it is unlike most of the other members of this family due to its propensity to cause persistent infections. This persistent infection eventually results in chronic liver disease, cirrhosis and hepatocellular carcinoma in a proportion of infected individuals. It has been difficult to examine correlates of clearance or persistence because most acute phase HCV infections are subclinical or result in symptoms which are non-specific; consequently, acute infections are not generally recognised and patients often present many years later with persistent infection and accompanying chronic liver disease. Nevertheless, seminal studies, performed during the acute phase, have identified a number of factors which are likely to influence the outcome of infection, although it is possible that the mechanism is multifactorial. One of these factors is impairment of dendritic cell function by a mechanism resulting from expression of an HCV protein(s) in these cells. This may be a major factor in the failure of the immune response to expand after HCV infection, leading to persistence. Nevertheless, it may be possible to overcome this defect by autologous transfusion of HCV antigen-loaded, mature dendritic cells and the purpose of this review is to highlight the need and general approaches for developing dendritic cell-based immunotherapy for HCV infection. PMID 15163415

Selective functional deficit in dendritic cell--T cell interaction is a crucial mechanism in chronic hepatitis B virus infection.
Apr. 2004 | Zheng, B J; Zhou, J; Qu, D; Siu, K L; Lam, T W; Lo, H Y; Lee, S S; Wen, Y M
A defect in specific T cell immunity has long been assumed to be the central mechanism of persistent Hepatitis B virus (HBV) infection. Recent studies on HBV transgenic mice have suggested, however, that functional deficit of dendritic cells (DC) was an underlying cause for the T cell dysfunction. The functions of monocyte-derived DC were determined by studying 75 subjects that included chronic hepatitis B patients with low or high HBV load; antibody to hepatitis B surface antigen (anti-HBs) positive individuals who had recovered completely from previous acute HBV infection; healthy donors who had received hepatitis B vaccination and were anti-HBs positive; and immunologically naïve to HBV or the vaccine individual. Impaired interactions between monocyte-derived DC and T cells were shown in chronic HBV infection patients, especially in those with active virus replication. The dysfunctions included: (i) failure of DC to increase human leukocyte antigen (HLA-II), B7 expression and interleukin-12 secretion in responses to hepatitis B surface antigen (HBsAg), (ii) defective induction of T cell proliferative response to HBsAg, (iii) failure to activate T cells to produce cytokines and (iv) deficit in the induction of antigen specific cytotoxic T lymphocytes (CTLs). In vitro treatment of DC with tumour necrosis factor-alpha improved HLA-II and B7 expression, as well as Th cell and CTL responses. It is concluded that defective DC-T cell interactions may account for the specific T cell immune defects in chronic HBV infection. Immunotherapy that aims at restoring DC functions could offer a new opportunity for effectively managing persistent HBV infections. PMID 15117323

Presence of functional dendritic cells in patients chronically infected with hepatitis C virus.
Jan. 2004 | Longman, Randy S; Talal, Andrew H; Jacobson, Ira M; Albert, Matthew L; Rice, Charles M
The absence of expanded numbers of hepatitis C virus (HCV)-reactive CD8(+) T lymphocytes (CTLs) in patients chronically infected with HCV has led to the investigation of dendritic cell (DC) function in this population as a potential cause for this defect. Several studies have shown evidence for impaired monocyte-derived DCs in chronically infected patients. As it is difficult to reconcile these data with the fact that patients with chronic HCV are immune competent, we re-evaluated this finding, carefully assessing phenotypic markers and functional activity of patient DCs as compared with noninfected controls. In contrast to these prior studies, DCs from 13 of 13 chronic HCV patients expressed typical maturation markers. These mature DCs were capable of priming allogeneic T lymphocytes, as well as stimulating influenza-specific memory T cells. This finding is consistent with clinical and immunologic data that the deficit in the patient's immune repertoire is HCV-specific and suggests that refined models are required for understanding the role of DCs in HCV pathogenesis. PMID 14525790

Hepatitis C virus structural proteins impair dendritic cell maturation and inhibit in vivo induction of cellular immune responses.
Sep. 2003 | Sarobe, Pablo; Lasarte, Juan José; Zabaleta, Aintzane; Arribillaga, Laura; Arina, Ainhoa; Melero, Ignacio; Borrás-Cuesta, Francisco; Prieto, Jesús
Hepatitis C virus (HCV) chronic infection is characterized by low or undetectable cellular immune responses against HCV antigens. Some studies have suggested that HCV proteins manipulate the immune system by suppressing the specific antiviral T-cell immunity. We have previously reported that the expression of HCV core and E1 proteins (CE1) in dendritic cells (DC) impairs their ability to prime T cells in vitro. We show here that immunization of mice with immature DC transduced with an adenovirus encoding HCV core and E1 antigens (AdCE1) induced lower CD4(+)- and CD8(+)-T-cell responses than immunization with DC transduced with an adenovirus encoding NS3 (AdNS3). However, no differences in the strength of the immune response were detected when animals were immunized with mature DC subsequently transduced with AdCE1 or AdNS3. According to these findings, we observed that the expression of CE1 in DC inhibited the maturation caused by tumor necrosis factor alpha or CD40L but not that induced by lipopolysaccharide. Blockade of DC maturation by CE1 was manifested by a lower expression of maturation surface markers and was associated with a reduced ability of AdCE1-transduced DC to activate CD4(+)- and CD8(+)-T-cell responses in vivo. Our results suggest that HCV CE1 proteins modulate T-cell responses by decreasing the stimulatory ability of DC in vivo via inhibition of their physiological maturation pathways. These findings are relevant for the design of therapeutic vaccination strategies in HCV-infected patients. PMID 14512536

Hepatitis B virus-induced defect of monocyte-derived dendritic cells leads to impaired T helper type 1 response in vitro: mechanisms for viral immune escape.
Juli 2003 | Beckebaum, Susanne; Cicinnati, Vito R; Zhang, Xia; Ferencik, Stanislav; Frilling, Andrea; Grosse-Wilde, Hans; Broelsch, Christoph Erich; Gerken, Guido
Dendritic cells (DC) are the most potent antigen-presenting cells and play a central role in the induction of antiviral immune responses. Recently, we have shown that monocyte-derived DC (MoDC) from patients with chronic hepatitis B virus (HBV) infection are functionally impaired. In our present study MoDC from healthy subjects were propagated in vitro and inoculated with HBV particles to investigate the precise mechanisms that underly MoDC dysfunction. T-cell proliferation assays revealed an impaired allostimulatory capacity of HBV-inoculated MoDC (HBV-MoDC) as well as a lower potential of stimulating autologous T cells against a recall antigen in comparison to control-MoDC. Interleukin-2, tumour necrosis factor-alpha and interferon-gamma production by T cells in proliferation assays with HBV-MoDC was significantly lower than with control-MoDC and correlated with lower IL-12 production in HBV-MoDC cultures. The presence of the nucleoside analogue lamivudine (3TC), an inhibitor of HBV replication, restored impaired allostimulatory function of HBV-MoDC and up-regulated major histocompatibility complex class II expression. These results show that HBV infection compromises the antigen-presenting function of MoDC with concomitant impairment of T helper cell type 1 responses. This may play an important role for viral immune escape leading to chronic HBV infection. However, 3TC treatment can overcome HBV-MoDC-related T-cell hyporeactivity and this underscores its important role in enhanced immune responses to HBV. PMID 12871214

Dendritic cell vaccination and viral infection--animal models.
Juni 2003 | Ludewig, B
Dendritic cells (DCs) play a pivotal role in the initiation and maintenance of immune responses against viruses and other microbial pathogens. Adoptively transferred, in vitro manipulated DCs presenting antigen derived from different viruses have been shown to elicit cytotoxic T cell (CTL) and T helper (Th) cell responses and to induce protective antiviral immunity. Furthermore, DC-based adoptive immunotherapies have the potential to specifically (re)activate antiviral immunity in chronic viral diseases such as HIV or hepatitis virus infections. Cellular dendritic cell vaccines, however, are not suitable for large-scale prophylactic immunization. Strategies for vaccine development should therefore aim at the specific delivery of microbial antigens to DCs in situ. Furthermore, appropriate mobilization and activation of DCs by the vaccine is important for the generation of optimal antimicrobial immune responses. Here, we discuss recent data on induction of antiviral immunity with various DC-vaccination approaches and outline future directions for the development of specific antigen targeting to DCs. PMID 12797449

Reduced virus specific T helper cell induction by autologous dendritic cells in patients with chronic hepatitis B - restoration by exogenous interleukin-12.
Sep. 2002 | Löhr, H F; Pingel, S; Böcher, W O; Bernhard, H; Herzog-Hauff, S; Rose-John, S; Galle, P R
Insufficient stimulatory capacities of autologous dendritic cells (DC) may contribute in part to impaired T cell stimulation and therefore viral persistence in patients with chronic hepatitis B virus (HBV) infection. In order to characterize the antigen presenting functions of DC from chronic HBV carriers and controls antigen specific T cell responses were analysed. CD34+ peripheral blood progenitor cells were differentiated to immature DC in the presence of GM-CSF, IL-6/IL-6R fusion protein and stem cell factor. Proliferative CD4+ T cell responses and specific cytokine release were analysed in co-cultures of DC pulsed with HBV surface and core antigens or tetanus toxoid and autologous CD4+ T cells. Cultured under identical conditions DC from chronic HBV carriers, individuals with acute resolved hepatitis B and healthy controls expressed similar phenotypical markers but chronic HBV carriers showed less frequent and weaker HBV antigen specific proliferative T helper cell responses and secreted less interferon-gamma while responses to the tetanus toxoid control antigen was not affected. Preincubation with recombinant IL-12 enhanced the HBV specific immune reactivities in chronic HBV patients and controls. In conclusion, the weak antiviral immune responses observed in chronic hepatitis B may result in part from insufficient T cell stimulating capacities of DC. Immunostimulation by IL-12 restored the HBV antigen specific T cell responses and could have some therapeutical benefit to overcome viral persistence. PMID 12296860

Activation and maturation of antigen-presenting dendritic cells during vaccine therapy in patients with chronic hepatitis due to hepatitis B virus.
Dez. 2001 | Horiike, Norio; Md Fazle Akbar, Sk; Ninomiya, Tuneyuki; Abe, Masanori; Michitaka, Kojiro; Onji, Morikazu
BACKGROUND/AIMS: The function and phenotype of antigen-presenting dendritic cells (DC) have been checked in patients with chronic hepatitis (CH) undergoing vaccine therapy to have insight about the mechanism of this immune therapy. SUBJECTS AND METHODS: Eleven patients with CH due to hepatitis B virus (HBV) were injected with vaccine containing 20 &mgr;g of hepatitis B surface antigen (HBsAg) once at every 2 weeks for 12 consecutive times. The serum interluekin-12 levels were estimated and the function and phenotype of peripheral blood DC were checked before, during and after the vaccine therapy. RESULTS: Sustained normalization of DNA-polymerase activity and alanine aminotransferase were seen in six patients (complete responders [CR]) due to vaccine therapy. The levels of serum interleukin-12 (P<0.01), the stimulatory capacity of peripheral blood DC (P<0.05) and the numbers of CD83-positive mature DC and CD86-positive activated DC (P<0.05) were significantly increased due to vaccination in CH-B patients, especially in younger patients. CONCLUSIONS: A role of DC during vaccine therapy is shown. This study also indicates that more effective vaccine therapy can be developed by upregulating the function of DC in situ in younger patients. PMID 12084554

Dendritic cells and chronic hepatitis virus carriers.
Aug. 2001 | Akbar, S M; Horiike, N; Onji, M; Hino, O
Many individuals infected with hepatitis B virus (HBV) and hepatitis C virus (HCV) are unable to clear these viruses following an acute infection and become chronically infected. There are more than 400 million HBV and HCV carriers in the world and a considerable number of these patients would eventually develop more severe complications like liver cirrhosis and hepatocellular carcinoma. It is not clearly known how an individual develops a chronic hepatitis virus carrier state; however, a defective immune response of the host is thought to play a critical role in the underlying pathogenetic mechanism. On the other hand, dendritic cells (DCs), the most potent antigen-presenting cells, are widely distributed in both lymphoid and nonlymphoid tissues. Recognition of the microbes or microbial antigens by DCs is one of critical events for the initiation of an immune response. DCs also play a cardinal role during the progression and termination of an immune response. The aim of this overview is to provide information regarding the role of DCs in the pathogenesis of chronic hepatitis due to HBV and HCV in humans and in animal models of HBV and HCV carrier states. First, we summarize our current understanding of the pathogenesis of hepatitis virus carrier states and also of general properties of DCs. Next, we discuss the data on the phenotypes and functions of DCs in both human and murine HBV and HCV carriers. We also discuss vaccine therapy in murine HBV carriers because activation of DCs due to vaccination-initiated HBsAg-specific immune responses in HBV transgenic mice (HBV-Tg), which in turn resulted in complete clearance of hepatitis B surface antigen and hepatitis B e antigen and decreased levels of HBV DNA in some HBV-Tg. Finally, we discuss the extracted questions and future research directions. PMID 11509880

Generation of hepatitis C virus-specific cytotoxic T lymphocytes from healthy individuals with peptide-pulsed dendritic cells.
Mai 2001 | Ito, A; Kanto, T; Kuzushita, N; Tatsumi, T; Sugimoto, Y; Miyagi, T; Takehara, T; Katayama, K; Mochizuki, K; Hiramatsu, N; Kasahara, A; Yoshiya, I; Sasaki, Y; Hori, M; Hayashi, N
In hepatitis C virus (HCV) infection, cytotoxic T lymphocytes (CTL) are involved in liver inflammation and contribute to the reduction of viral load. Antibodies for HCV-CTL precursor frequencies (CTLpf) are relatively low in chronic hepatitis C, and this may be related to the poor CTL response in vivo. The aim of this study was to assess the efficacy of dendritic cells (DC) as antigen-presenting cells in CTL generation from low CTLpf. PMID 11339423

Dendritic cell immunization breaks cytotoxic T lymphocyte tolerance in hepatitis B virus transgenic mice.
Nov. 1998 | Shimizu, Y; Guidotti, L G; Fowler, P; Chisari, F V
Hepatitis B virus (HBV) transgenic mice that are immunologically tolerant to HBV-encoded Ags represent a model of chronic HBV infection suitable for the development of therapeutic immunization strategies before testing in humans. Five lineages of HBV transgenic mice were immunized with plasmid DNA that encodes hepatitis B surface Ag (HBsAg) or with cytokine-activated bone marrow-derived dendritic cells (DCs) in an attempt to break tolerance to HBsAg at the B and T cell levels. DNA immunization stimulated an Ab response but not a cytotoxic T lymphocyte response to HBsAg in two of the five transgenic lineages studied. In contrast, infusion of activated transgenic or nontransgenic DCs stimulated a splenic CTL response in all three transgenic lineages immunized in this manner at precursor frequencies comparable to those in nontransgenic mice, indicating that DC function is normal, and HBsAg-specific CTLs are present but functionally silent in these transgenic animals. Importantly, none of the animals developed hepatitis or displayed suppressed viral gene expression or replication following either DNA immunization or DC administration even in the presence of anti-hepatitis B surface (HBs) Abs and HBs-specific CTLs. These results indicate that Ag presentation by cytokine-activated DCs can break tolerance and trigger an anti-viral CTL response in HBV transgenic mice, and they suggest that this strategy is more efficient than DNA immunization in this setting. Nonetheless, more efficient immunization strategies are needed to stimulate an immune response of sufficient quality and magnitude to achieve an immunotherapeutic antiviral effect. PMID 9794377

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