Biomedical subjects

Heribert Stoiber

Publications and source records attributed to Heribert Stoiber.

At least 19 recordsLinked to original sources

PubMed2007

Tracing complement-retroviral interactions from mucosal surfaces to the lymphatic tissue.

During (nearly) all steps in retroviral pathogenesis, viruses are confronted with complement and complement receptor (CR)-positive cells. As all of the retroviruses tested so far activate the complement system, members of this virus family have adapted different protection mechanisms to keep complement activation under the threshold necessary to avoid complement-mediated lysis. As a consequence of complement activation, retroviruses are covered with complement proteins and thus provide additional ligands to interact with CR-expressing cells. This review discusses the complex complement-retroviral interactions and follows the fate of the virus on its way to the lymphatic tissue.

Animals↗

PubMed2007

A single vaccination with attenuated SIVmac 239 via the tonsillar route confers partial protection against challenge with SIVmac 251 at a distant mucosal site, the rectum.

Elucidating the mechanisms that protect monkeys previously immunized with attenuated SIV (SIVDeltanef) against challenge infection with pathogenic virus may reveal new strategies for the development of an effective HIV vaccine. Here we show that a single atraumatic application of SIVDeltanef to the tonsils of four rhesus macaques conferred protection against SIVmac251 applied intrarectally 26 weeks later. While this protection was not complete, i.e., challenge virus could be isolated from all immunized animals, it was reflected by significantly lower viral loads in the blood (weeks 2-16 after challenge, p < 0.01) and considerably lower loads in lymphoid organs, and more stable peripheral CD4 counts in a proportion of the immunized animals as compared to four non-immunized, SIVmac251-infected control monkeys. SIV-specific humoral as well as systemic and mucosal T cell responses were detected in the immunized animals, but there was no correlation between their magnitude of expression and the level of protection. Analyses of leukocyte subsets in these animals at necropsy (24 weeks after challenge) did not reveal a significantly enhanced proportion of gamma/delta T cells in the tissues of protected monkeys. Therefore, tonsillar application of attenuated SIV induces protection in some animals against a superinfection with wild-type SIV distant at a distant mucosal site.

Animals↗

PubMed2007

Float on: lipid rafts in the lifecycle of HIV.

All steps in the HIV lifecycle--entry, assembly, budding, induction of signal transduction and subsequent cell activation--are complex multifactorial mechanisms where cholesterol and sphingolipids (glycosphingolipids--GSLs and sphingomyelins in mammalian cells) are closely involved. Here we will review the contribution of these heterogeneous membrane lipid microdomains, referred to as lipid rafts, DIGs (detergent-insoluble glycolipid-enriched complexes), DRMs (detergent-resistant membranes), GEMs (glycolipid-enriched membranes), caveolae, TIMs (Triton-insoluble membranes) for interactions of HIV with the host cell. The accurate terminology was discussed elsewhere, and to simplify matters we will use rafts or lipid rafts throughout the review.

Gene Products, nef↗

PubMed2007

Changes in HIV-specific antibody responses and neutralization titers in patients under ART.

In this study, we tested for antibody reactivities against gp120 and gp41-derived peptides, recombinant gp160, gp41 and tat in HIV-positive sera under antiretroviral therapy (ART) and determined their neutralization capacity. As a baseline, sera from patients in stage A, B and C of the disease, long term non-progressors (LNPs) and HIV-negative individuals were included. Compared to LNPs or sera from patients in group A, the reactivity of sera in stage B or C against gp120-derived peptides was reduced parallel to disease progression. Reactivity of these samples was compared with sera of patients under ART. Parallel to the decrease of viral load, the reactivity against gp120 and gp41-derived epitopes, recombinant gp160 and gp41 or the native gp120/41 complex was significantly reduced. Antibody-mediated neutralization of HIV-1 was detectable prior to ART but revealed substantial decreases coupled with progression of therapy. Responses to recombinant tat dropped after three months of therapy, increased however at later time points to initial levels. These data indicate that in parallel to the decrease in viral load and antibodies against gp120, the neutralization capacity of sera under ART is reduced, and can not be compensated by an increase in tat-specific antibodies.

Antibodies, Viral↗

PubMed2006

Factor I-mediated processing of complement fragments on HIV immune complexes targets HIV to CR2-expressing B cells and facilitates B cell-mediated transmission of opsonized HIV to T cells.

Our study demonstrates that binding of complement-opsonized HIV to complement receptor type 1 on human erythrocytes (E) via C3b fragments is followed by a rapid normal human serum-mediated detachment of HIV from E. The release was dependent on the presence of factor I indicating a conversion of C3b fragments to iC3b and C3d on the viral surface. This in turn resulted in an efficient binding of opsonized HIV to CR2-expressing B cells, thus facilitating B cell-mediated transmission of HIV to T cells. These data provide a new dynamic view of complement opsonization of HIV, suggesting that association of virus with E might be a transient phenomenon and the factor I-mediated processing of C3b to iC3b and C3d on HIV targets the virus to complement receptor type 2-expressing cells. Thus, factor I in concert with CR1 on E and factor H in serum due to their cofactor activity are likely to be important contributors for the generation of C3d-opsonized infectious HIV reservoirs on follicular dendritic cells and/or B cells in HIV-infected individuals.

B-Lymphocytes↗

PubMed2006

Immunogenicity and efficacy of immunodeficiency virus-like particles pseudotyped with the G protein of vesicular stomatitis virus.

Vaccination with exogenous antigens such as recombinant viral proteins, immunodeficiency virus-derived whole inactivated virus particles, or virus-like particles (VLP) has generally failed to provide sufficient protection in animal models for AIDS. Pseudotyping VLPs with the vesicular stomatitis virus G protein (VSV-G), which is known to mediate entry into dendritic cells, might allow more efficient stimulation of immune responses. Therefore, we pseudotyped noninfectious immunodeficiency virus-like particles with VSV-G and carried out a preliminary screen of their immunogenicity and vaccination efficacy. Incorporation of VSV-G into HIV-1 VLPs led to hundred-fold higher antibody titers to HIV-1 Gag and enhancement of T cell responses in mice. Repeated vaccination of rhesus monkeys for 65 weeks with VSV-G pseudotyped simian immunodeficiency virus (SIV)-like particles (VLP[G]) provided initial evidence for efficient suppression of viral load after mucosal challenge with the SIVmac239 virus. Challenge of monkeys after a 28 week vaccination regimen with VLP[G] led to a reduction in peak viremia, but persistent suppression of viral load was not achieved. Due to limitations in the number of animals available for this study, improved efficacy of VSV-G pseudotyped VLPs in nonhuman primates could not be demonstrated. However, mouse experiments revealed that pseudotyping of VLPs with fusion-competent VSV-G clearly improves their immunogenicity. Additional strategies, particularly adjuvants, should be considered to provide greater protection against a challenge with pathogenic immunodeficiency virus.

AIDS Vaccines↗

PubMed2005

Reduction of viral loads by multigenic DNA priming and adenovirus boosting in the SIVmac-macaque model.

In this study, we investigated the ability of a multigenic SIV DNA prime/replication-defective adenovirus serotype 5 (rAd/SIV) boost regimen to induce SIV-specific immune responses and protection against intrarectal challenge with SIVmac251 in rhesus macaques. Four rhesus macaques were immunized intramuscularly three times at 8-week intervals with SIV DNA vaccine and boosted once with rAd/SIV vaccine Four control macaques received the same amount of mock plasmid DNA and mock adenovirus vector. While the SIV DNA vaccine included plasmids expressing a mutated human IL-12 gene (IL-12N222L) as well as SIVmac239 structural and regulatory genes, the rAd/SIV vaccine contained rAd vectors expressing SIVmac239 genes only. Immunization with SIV DNA vaccine alone induced SIV-specific IFN-gamma ELISPOT responses in only two of four vaccinated macaques, whereas all animals developed SIV-specific T-cell responses and Env- and Tat-specific antibody responses following the rAd/SIV vaccine boost. Upon intrarectal challenge with pathogenic SIVmac251, strong anamnestic Env-specific binding and neutralizing antibody responses were detected in the vaccinated macaques. Overall, the immunized macaques had lower peak and set-point viral loads than control macaques, suggesting that the induced immune responses play a role in the control of viremia. In addition, the loss of CD4+ T cells was delayed in the vaccinated macaques after challenge. These results indicate that the multigenic DNA prime-adenovirus boost immunization may be a promising approach in developing an effective AIDS vaccine.

Adenoviridae↗

PubMed2005

Complement dependent trapping of infectious HIV in human lymphoid tissues.

OBJECTIVES: HIV-1 bound extracellularly to follicular dendritic cells (FDC) in germinal centers (GC) of lymphoid tissues (LT) represents the largest viral reservoir in HIV-infected individuals; however there is no direct evidence as to whether HIV trapped in human GC remains infectious. In the GC, complement receptors and Fc gamma receptors have been suggested to participate in trapping of HIV; therefore, the relative contribution of complement- and Fc gamma receptors in binding HIV on LT was investigated and the infectivity of this virus was tested. DESIGN: As it is difficult to isolate FDC without contaminations of productively infected cells, HIV was detached from LT of HIV positive individuals using antibodies blocking complement- and Fc gamma receptors. Isolated virus was tested in an infectivity assay. METHODS: HIV detached from LT was quantified by HIV p24 ELISA, PCR and in an in vitro infectivity assay. Presence and accessibility of viral envelope proteins, complement factors and immunoglobulins on the surface of detached viral particles were evaluated through viral capture by respective antibodies. RESULTS: Although both C3d-fragments and IgG molecules were identified on the surface of HIV detached from LT, trapping of HIV was mediated solely by CR2-C3d interactions, whereas contribution of Fc gamma receptors was not detectable. Infectivity assays with HIV stripped from LT of HIV positive individuals revealed that in four out of ten patients HIV particles were infectious. CONCLUSIONS: These findings indicate that in the GC infectious virus is trapped on CR2-expressing FDC (or B cells). Reduction of this pool of HIV could be a therapeutic goal.

Cells, Cultured↗

PubMed2005

C-type lectin-independent interaction of complement opsonized HIV with monocyte-derived dendritic cells.

HIV directly activates the complement cascade and is, therefore, opsonized with C3-cleavage products in vivo. This cloud of C3 fragments on the viral surface may impair the interaction of the HIV envelope glycoproteins gp120/gp41 with C-type lectins expressed on immature dendritic cells (iDC). Therefore, we determined the accessibility of gp120 after opsonization and compared the interaction of DC with non-opsonized or complement-opsonized HIV. The recognition of native gp120 was drastically impaired when the virus was covered by complement. Independent of opsonization, similar amounts of HIV bound to DC. The interaction of iDC and the infection of DC-PBL co-cultures with non-opsonized virus was significantly reduced by mannan and antibodies which inhibit the ICAM-1-CR3 interaction. The binding of opsonized virus to iDC was reduced by an anti-CR3-antibody, which interferes with the binding of C3 fragments, but was not affected by mannan. Complement enhanced the HIV infection of DC and DC-PBL co-cultures significantly. Mannan did not inhibit the complement-dependent enhancement of infection. Thus, non-opsonized and opsonized HIV interacted with iDC, although the binding mechanisms seemed to differ. As HIV is opsonized in vivo, the C-type lectin-independent interaction of opsonized viruses with iDC has to be taken into account.

Antibodies, Monoclonal↗

PubMed2005

Mechanisms promoting dendritic cell-mediated transmission of HIV.

Dendritic cells (DC) survey epithelial or mucosal surfaces for antigens, take them up via their endocytic or phagocytic receptors, process the captured antigens and migrate to the lymphatic tissues. In the draining lymph nodes they present the immunogenic peptides to T cells thereby inducing antigen-specific immune responses. HIV-1 in turn seems to have developed mechanisms to exploit the natural trafficking of DC to establish infection in its primary targets, the CD4+ T cells. This review discusses several aspects of DC-HIV interactions with a main focus on the attachment, internalisation and transmission of the virus by DC to cells, susceptible for infection with HIV.

Dendritic Cells↗

PubMed2005

Complement-opsonized HIV: the free rider on its way to infection.

The complement system (C) is one of the main humoral components of innate immunity. Three major tasks of C against invading pathogens are: (i) lysis of pathogens by the formation of the membrane attack complex (MAC); (ii) opsonization of pathogens with complement fragments to favor phagocytosis; and (iii) attraction of inflammatory cells by chemotaxis. Like other particles, HIV activates C and becomes opsonized. To escape complement-mediated lysis, HIV has adopted various properties, which include the acquisition of HIV-associated molecules (HAMs) belonging to the family of complement regulators, such as CD46, CD55, CD59, and the interaction with humoral regulatory factors like factor H (fH). Opsonized virus may bind to complement receptor positive cells to infect them more efficiently or to remain bound on the surface of such cells. In the latter case HIV can be transmitted to cells susceptible for infection. This review discusses several aspects of C-HIV interactions and provides a model for the dynamics of this process.

Complement Inactivator Proteins↗

PubMed2004

HIV-1-induced migration of monocyte-derived dendritic cells is associated with differential activation of MAPK pathways.

From the site of transmission at mucosal surfaces, HIV is thought to be transported by DCs to lymphoid tissues. To initiate migration, HIV needs to activate DCs. This activation, reflected by intra- and extracellular changes in cell phenotype, is investigated in the present study. In two-thirds of the donors, R5- and X4-tropic HIV-1 strains induced partial up-regulation of DC activation markers such as CD83 and CD86. In addition, CCR7 expression was increased. HIV-1 initiated a transient phosphorylation of p44/p42 ERK1/2 in iDCs, whereas p38 MAPK was activated in both iDCs and mDCs. Up-regulation of CD83 and CD86 on DCs was blocked when cells were incubated with specific p38 MAPK inhibitors before HIV-1-addition. CCR7 expression induced by HIV-1 was sufficient to initiate migration of DCs in the presence of secondary lymphoid tissue chemokine (CCL21) and MIP-3beta (CCL19). Preincubation of DCs with a p38 MAPK inhibitor blocked CCR7-dependent DC migration. Migrating DCs were able to induce infection of autologous unstimulated PBLs in the Transwell system. These data indicate that HIV-1 triggers a cell-specific signaling machinery, thereby manipulating DCs to migrate along a chemokine gradient, which results in productive infection of nonstimulated CD4(+) cells.

Antigens, Differentiation, Myelomonocytic↗

PubMed2004

HIV and human complement: inefficient virolysis and effective adherence.

Both, HIV envelope proteins gp120 and gp41 can directly activate complement system, even in the absence of HIV-specific antibodies. During the budding process HIV acquires host membrane-associated molecules among these complement regulatory proteins (CRPs). The presence of CRPs on the viral surface rescues HIV from complement-mediated virolysis. The inefficient virolysis results in the deposition of complement-fragments on the viral surface allowing interactions of HIV with complement receptor expressing cells. In this review, the interaction of HIV with the complement system and the consequences of complement opsonisation on virus infection will be discussed.

Acquired Immunodeficiency Syndrome↗

PubMed2004

Passive immunization with the anti-HIV-1 human monoclonal antibody (hMAb) 4E10 and the hMAb combination 4E10/2F5/2G12.

OBJECTIVES: To study the safety, immunogenicity and pharmacokinetics of the human monoclonal antibody (hMAb) 4E10 alone and in combination with the hMAbs 2F5 and 2G12 in HIV-1-infected persons. MATERIALS AND METHODS: Eight healthy volunteers with > or =350 CD4 cells/mm3 and < or =100 000 HIV-1 RNA copies/mL were enrolled, seven finished the study. A single 4E10 infusion was administered on day 0, followed by three doses of the hMAb combination 4E10/2F5/2G12 on days 7, 14 and 21 (total amount 8.5 g). Safety was assessed by physical examination, blood chemistry, complete blood cell count and recording of adverse events. 4E10, 2F5 and 2G12 plasma levels were determined before and at the end of each infusion and during the 7 week follow-up. RESULTS: No drug-related adverse events were observed throughout the study. The median plasma concentrations immediately after the first infusion were 371, 253 and 139 microg/mL for 4E10, 2F5 and 2G12. Multiple infusions resulted in maximum plasma concentrations of 407, 294 and 210 microg/mL for 4E10, 2F5, and 2G12, respectively. The median elimination half-lives (t1/2beta) were 6.6, 3.2 and 14.1 days for 4E10, 2F5 and 2G12. A low level antibody response against 2G12 was found in two patients. CONCLUSION: This Phase I trial showed that the hMAb 4E10 can be safely administered, both alone and in combination with 2F5 and 2G12 to HIV-1-infected patients.

Acquired Immunodeficiency Syndrome↗

PubMed2004

Early protection against pathogenic virus infection at a mucosal challenge site after vaccination with attenuated simian immunodeficiency virus.

Atraumatic application of attenuated SIVmac239 Delta nef vaccine to the tonsils of rhesus macaques provided protection against challenge 26 weeks later with infectious SIVmac251 applied through this route. Early events at the mucosal portal of entry of challenge virus were followed. Wild-type virus was detected in nonvaccinated controls by day 4, and then simian immunodeficiency virus (SIV) replicated vigorously at days 7 and 14. In contrast, a challenge of 10 of 10 vaccinees with SIV did not significantly raise RNA levels in the plasma or increase infected cells in lymphoid tissues, as assessed by single-cell labeling for viral RNA and nef protein. Vaccine virus was found in the tonsils of all vaccinees, but challenge virus was only detected at this portal of entry in 4 of 10 monkeys. In the tonsil, the challenge virus did not induce an expansion of perforin(+) killer cells. However, there was a significant increase in gamma delta T cells and mature dendritic cells relative to unvaccinated controls. Therefore, during tonsillar SIV Delta nef vaccination, infection is blocked early at the entry portal, which we propose is due in part to innate functions of gamma delta T and dendritic cells.

Animals↗

PubMed2004

Complement synthesis and activation in the brain of SIV-infected monkeys.

Complement is one of the most critical defence tools against cerebral infections, but uncontrolled complement biosynthesis and activation can induce profound brain tissue damage. To clarify the role of complement in the pathogenesis of AIDS-associated neurological disorders, we analysed the synthesis of complement in the brains of SIV-infected rhesus macaques. Using immunohistochemical staining we could show that the cerebral synthesis of complement factors C1q and C3 was strongly upregulated in SIV-infected monkeys compared to the spontaneous synthesis in uninfected control monkeys. Astrocytes, neurons, microglia, infiltrating macrophages and multinuclear giant cells all contribute to the high amounts of C1q and C3 in the brain. Secreted C1q and C3 are also deposited on the membrane of neurons, a prerequisite for formation of the membrane-driven lytic membrane attack complex. The membrane deposition thus might suggest complement-induced lysis of bystander neurons as a potential mechanism for cell damage during viral infection of the brain.

Animals↗

PubMed2004

HIV-1 induces complement factor C3 synthesis in astrocytes and neurons by modulation of promoter activity.

Virus-induced complement expression and activation in the brain is hypothesized to contribute to the process of neurodegeneration in AIDS-associated neurological disorders. Previous experiments have shown that the human immunodeficiency virus (HIV) upregulates the low basal production of complement factor C3 in astrocytes and neurons. Since inhibition of complement synthesis and activation in the brain may represent a putative therapeutic goal to prevent virus-induced damage, we analysed the mechanism of the HIV-induced modulation of C3 expression. Detailed studies using different C3 promoter constructs revealed that HIV activates the synthesis of C3 by stimulation of the promoter. This HIV-induced promoter activation could be measured both in different astrocytic cell lines and in neurons. Deletion constructs of the C3 promoter defined the IL-6/IL-1beta responsive element within the promoter region as a central element for the responsiveness of the C3 promoter towards the influence of HIV. A binding site for the transcription factor C/EBPdelta was identified as important regulatory domain within the IL-6/IL-1beta responsive element, since a point mutation which eliminates the binding capacity of C/EBPdelta to this site also abolishes the induction by HIV-1. Similarly, the viral proteins Nef and gp41 which had also been shown to stimulate the synthesis of C3, exert their effect via the IL-6/IL-1beta responsive element with binding of the transcription factor C/EBPdelta representing the critical step. Our experiments clearly define the mechanism for the induction of complement factors in the HIV-infected brain and reveal a decisive role of the regulator protein C/EBPdelta for the HIV-induced increase in C3 expression.

Astrocytes↗

PubMed2004

Activation of mitogen activated protein kinases via complement receptor type 2.

BACKGROUND: Complement receptor type 2 (CR2) is the receptor for C3d and C3dg and for Epstein-Barr virus. The aim of our study was to explore whether CR2 can independently mediate the activation of mitogen-activated protein kinases (MAPKs, including ERK, JNK, and p38MAPK), and to highlight the molecular mechanism of CD4+ cell deletion in AIDS. METHODS: HOS cells (HOS-CR2) and HOS-CD4 cells (HOS-CD4CR2) stably expressing CR2 were established and then identified by FACS and Western blotting. Activation and blocking tests of MAPKs were assessed by Western blot. Cell proliferation was determined using Cell Titer 96((R)) Aqueous One Solution Reagent. RESULTS: FACS results showed that the positive rates of HOS-CR2 and HOS-CD4CR2 cells were greater than 96%, and Western blot showed that the CR2 expression levels on HOS-CR2 and HOS-CD4CR2 cells were high. Activation and blocking tests of MAPKs (ERK, JNK, and p38MAPK) were carried out in HOS-CR2, HOS-CD4, and HOS-CD4CR2 cells. The activation of MAPKs in HOS-CR2 cells stimulated with PMA (100 ng/ml) and NHS (10%) was identical. The activation of MAPKs increased at 5 minutes, reached a peak at 10 minutes, and decreased to baseline within 30 minutes, all in a time-dependent manner; the activation of MAPKs was blocked by anti-CR2 McAb, PD98059 (inhibitor of ERK), and Wortmanin (inhibitor of PI-3K), respectively. In HOS-CD4 cells, MAPKs were activated by HIV-gp160. In HOS-CD4CR2 cells, MAPK activation was induced by HIV-gp160, 10% NHS, and HIV-gp160 + 10% NHS; phosphorylation of p38MAPK was dramatically induced by HIV-gp160 + NHS, and lasted for 1 hour. The cell proliferation results showed that HIV-gp160 inhibited the proliferation of HOS-CD4 and HOS-CD4CR2 cells (P < 0.01) and that NHS enhanced the effect of HIV-gp160 (P < 0.01). CONCLUSIONS: The activation of MAPKs is independently mediated by CR2 and that anti-CR2 McAb, PD98059, and Wortmanin block the activation of MAPKs, respectively. The results of the signal transduction and cell proliferation assays of HOS-CD4CR2 cells show that CR2 plays a role in the pathogenesis of HIV infection, especially in the inhibition of CD4+ cell proliferation.

Cell Division↗