DC-SIGN is a receptor for human herpesvirus 8 on dendritic cells and macrophages

Human herpesvirus 8 (HHV-8) causes Kaposi's sarcoma and pleural effusion lymphoma. In this study, we show that dendritic cell-specific ICAM-3 grabbing nonintegrin (DC-SIGN; CD209) is a receptor for HHV-8 infection of myeloid DCs and macrophages. DC-SIGN was required for virus attachment to these cells and DC-SIGN-expressing cell lines. HHV-8 binding and infection were blocked by anti-DC-SIGN mAb and soluble DC-SIGN, and mannan, a natural ligand for DC-SIGN. Infection of DCs and macrophages with HHV-8 led to production of viral proteins, with little production of viral DNA, similar to HHV-8 infection of vascular endothelial cells. Infection of DCs resulted in down-regulation of DC-SIGN, a decrease in endocytic activity, and an inhibition of Ag stimulation of CD8+ T cells. We propose that DC-SIGN serves as a portal for immune dysfunction and oncogenesis caused by HHV-8 infection.     

Human herpesvirus 8 (HHV-8/KSHV) and hematologic malignancies

Human herpesvirus 8 (HHV-8), also defined Kaposi's sarcoma (KS)-associated herpesvirus, was identified by Chang and colleagues in 1994 using purely molecular techniques, before any serological evidence or virus isolation in cell culture could be achieved. HHV-8 is unique among herpesviruses because its prevalence in the general population is low and because it possesses the richest weaponry of viral oncogenes and tumor-promoting factors ever described. Eleven HHV-8-specific genes are homologs of cellular genes, which were hijacked from the host during a long parallel evolution, and at least five of such genes show both in vitro and in vivo transforming ability. HHV-8 is the causative agent of KS, but it has also been associated with different hematologic malignancies, including primary effusion lymphoma (PEL), multicentric Castelman's disease (MCD), MCD-related immunoblastic/plasmablastic lymphoma and various atypical lymphoproliferative disorders. Although low-level silent infection was detected in bone marrow stromal cells from patients with multiple myeloma, a role of HHV-8 in this disease is unlikely. As seen with KS, the incidence of HHV-8-associated lymphoproliferative disorders is increased in the setting of human immunodeficiency virus infection.     

Human herpesvirus 8 infects and replicates in primary cultures of activated B lymphocytes through DC-SIGN

Human herpesvirus 8 (HHV-8) is the etiological agent of Kaposi's sarcoma, primary effusion lymphoma, and some forms of multicentric Castleman's disease. Although latent HHV-8 DNA can be detected in B cells from persons with these cancers, there is little information on the replication of HHV-8 in B cells. Indeed, B cells are relatively resistant to HHV-8 infection in vitro. We have recently shown that DC-SIGN, a C-type lectin first identified on dendritic cells (DC), is an entry receptor for HHV-8 on DC and macrophages. We have also demonstrated previously that B lymphocytes from peripheral blood and tonsils express DC-SIGN and that this expression increases after B-cell activation. Here we show that activated blood and tonsillar B cells can be productively infected with HHV-8, as measured by an increase in viral DNA, the expression of viral lytic and latency proteins, and the production of infectious virus. The infection of B cells with HHV-8 was blocked by the pretreatment of the cells with antibody specific for DC-SIGN or with mannan but not antibody specific for xCT, a cystine/glutamate exchange transporter that has been implicated in HHV-8 fusion to cells. The infection of B cells with HHV-8 resulted in increased expression of DC-SIGN and a decrease in the expression of CD20 and major histocompatibility complex class I. HHV-8 could also infect and replicate in B-cell lines transduced to express full-length DC-SIGN but not in B-cell lines transduced to express DC-SIGN lacking the transmembrane domain, demonstrating that the entry of HHV-8 into B cells is related to DC-SIGN-mediated endocytosis. The role of endocytosis in viral entry into activated B cells was confirmed by blocking HHV-8 infection with endocytic pathway inhibitors. Thus, the expression of DC-SIGN is essential for productive HHV-8 infection of and replication in B cells.     

Characterization of monoclonal antibodies raised against the latent nuclear antigen of human herpesvirus 8

Human herpesvirus 8 (HHV-8; also designated Kaposi's sarcoma-associated herpesvirus) is the likely etiological agent of Kaposi's sarcoma (KS). HHV-8 encodes a latent nuclear antigen (LNA) which is the product of the viral gene orf 73. LNA is recognized by most infected patient sera and is the basis of current immunofluorescence assays used in epidemiological studies of HHV-8 infection. Here we describe the characterization of four monoclonal antibodies raised to the C-terminal third of LNA-glutathione S-transferase fusion proteins. These monoclonal antibodies recognized discrete linear epitopes within the C terminus and repetitive region of LNA, detected antigen in primary effusion lymphoma (PEL) cells, and precipitated a 220- to 230-kDa protein doublet corresponding to LNA from HHV-8-infected PEL cell lines. In situ immunocytochemistry of KS lesions with these antibodies show that LNA is extensively expressed in KS spindle cells.     

Human herpesvirus 8: an update

The discovery of human herpesvirus 8 (HHV-8) has opened a new field in the biology and in the clinical definition of HHV-8-associated diseases. Various aspects of epidemiology, laboratory diagnosis and treatment of HHV-8 infection are still controversial. This review will summarise the most recent findings on the modalities of viral transmission in geographic areas where the virus is endemic as compared with those where the infection is sporadic. The studies on the interactions between HHV-8 and Epstein-Barr virus in the promotion of tumorigenesis will be then emphasised. This review will finally address the problems still open in the laboratory diagnosis and in the definition of antiviral drug susceptibility of HHV-8. Solving satisfactorily these issues will be necessary to better understand the epidemiology of HHV-8 infection, to reduce the risk of viral transmission, the progression of infected hosts to HHV-8-related malignancies and to achieve more active treatment options for these clinical conditions.     

Characterization of entry mechanisms of human herpesvirus 8 by using an Rta-dependent reporter cell line

To analyze the mechanisms of entry of human herpesvirus 8 (HHV-8), we established a reporter cell line T1H6 that contains the lacZ gene under the control of the polyadenylated nuclear RNA promoter, known to be strongly activated by a viral transactivator, Rta. We found that infection with cell-free virus, as well as cocultivation with HHV-8-positive primary effusion lymphoma cell lines, activated the lacZ gene of T1H6 in a sensitive and dose-dependent manner. Addition of Polybrene and centrifugation enhanced, but polysulfonate compounds inhibited, the HHV-8 infectivity. RGD-motif-containing polypeptides and integrins did not decrease the infectivity, suggesting the presence of an additional cellular receptor other than the reported one. The entry was dependent on pH acidification but not on the clathrin pathway. Although conditioned media obtained from human immunodeficiency virus (HIV)-infected cells did not have any effect on the early steps of HHV-8 infection, intracellular expression of a proviral HIV type 1, but not of Tat alone, increased the HHV-8-dependent reporter activation slightly, suggesting a potential of HIV-mediated enhancement of an early step of HHV-8 infection.     

Gamma delta+ T cells involvement in viral immune control of chronic human herpesvirus 8 infection

Little is known about what effector populations are associated with the control of human herpesvirus 8 (HHV-8) infection in vivo. We compared T lymphocyte subsets among HIV-HHV-8+ and HIV-HHV-8- infected human individuals. alphabeta+ T cells from HHV-8-infected individuals displayed a significantly higher percentage of differentiated effector cells among both CD4+ and CD8+ T cell subsets. HHV-8 infection was associated with significant expansion of gammadelta+ Vdelta1 T cells expressing a differentiated effector cell phenotype in peripheral blood. In vitro stimulation of PBMC from HHV-8-infected individuals with either infectious viral particles or different HHV-8 viral proteins resulted in gammadelta Vdelta1 T cell activation. In addition, gammadelta Vdelta1 T cells displayed a strong reactivity against HHV-8-infected cell lines and prevented the release of infectious viral particles following the induction of lyric replication. These data indicate that gammadelta T cells play a role in both innate and adaptive T cell responses against HHV-8 in immunocompetent individuals.     

Kaposi's sarcoma-associated herpesvirus/human herpesvirus 8 RTA reactivates murine gammaherpesvirus 68 from latency

Murine gammaherpesvirus 68 (MHV-68), Kaposi's sarcoma-associated herpesvirus (HHV-8), and Epstein-Barr virus (EBV) are all members of the gammaherpesvirus family, characterized by their ability to establish latency in lymphocytes. The RTA protein, conserved in all gammaherpesviruses, is known to play a critical role in reactivation from latency. Here we report that HHV-8 RTA, not EBV RTA, was able to induce MHV-68 lytic viral proteins and DNA replication and processing and produce viable MHV-68 virions from latently infected cells at levels similar to those for MHV-68 RTA. HHV-8 RTA was also able to activate two MHV-68 lytic promoters, whereas EBV RTA was not. In order to define the domains of RTA responsible for their functional differences in viral promoter activation and initiation of the MHV-68 lytic cycle, chimeric RTA proteins were constructed by exchanging the N-terminal and C-terminal domains of the RTA proteins. Our data suggest that the species specificity of MHV-68 RTA resides in the N-terminal DNA binding domain.     

人类8型疱疹病毒的研究进展

人类8型疱疹病毒（human herpesvirus-8,HHV-8）又称卡波氏肉瘤相关疱疹病毒（Kapo- si's sarcoma- associated herpesvirus,KSHV）,是一种新的肿瘤病毒,目前被认为是卡波氏肉瘤（Kaposi's sarcoma,KS）致病因子,并且与primary effusion lymphoma （PEL）和multicentric Castleman's disease（MCD）相关。该病毒编码许多蛋白,包括潜伏感染相关蛋白,裂解感染相关蛋白和HHV-8特有基因表达蛋白,在KS和HHV-8相关疾病的发病中起到关键作用。     

Non-detection of human herpesvirus 8 (HHV-8) DNA in HHV-8-seropositive blood donors from three Brazilian regions

Human herpesvirus 8 (HHV-8), also known as Kaposi''s sarcoma-associated herpesvirus (KSHV), is the etiologic agent of all forms of Kaposi''s sarcoma, primary effusion lymphoma and the plasmablastic cell variant of multicentric Castleman disease. In endemic areas of sub-Saharan Africa, blood transfusions have been associated with a substantial risk of HHV-8 transmission. By contrast, several studies among healthy blood donors from North America have failed to detect HHV-8 DNA in samples of seropositive individuals. In this study, using a real-time PCR assay, we investigated the presence of HHV-8 DNA in whole-blood samples of 803 HHV-8 blood donors from three Brazilian states (São Paulo, Amazon, Bahia) who tested positive for HHV-8 antibodies, in a previous multicenter study. HHV-8 DNA was not detected in any sample. Our findings do not support the introduction of routine HHV-8 screening among healthy blood donors in Brazil. (WC = 140).     

Multiplex real-time PCR for the simultaneous detection of herpes simplex virus, human herpesvirus 6, and human herpesvirus 7

A simultaneous detection system to quantify HSV, HHV-6, and HHV-7 DNA via multiplex real-time PCR using different fluorochromes was developed. The minimum quantitative level established via this multiplex assay was four copies per reaction for HSV type 1, four copies for HHV-6, and three copies for HHV-7, respectively. The dynamic range encompassed at least six orders of magnitude. The system was specific and reproducible even in the presence of large amounts of other viral DNA. We then applied this multiplex real-time PCR assay to 105 CSF specimens obtained from subjects less than 15 years old in whom a diagnosis of viral encephalitis/encephalopathy was suspected on clinical grounds. The detection rate for each viral DNA was 6.7% for HSV, 9.5% for HHV-6, and 1.9% for HHV-7. These results indicate that our system is reliable and may be useful for the rapid diagnosis of viral encephalitis/encephalopathy.