No evidence for consistent virus-specific immunity in simian immunodeficiency virus-exposed, uninfected rhesus monkeys

Defining the immune correlates of the protection against human immunodeficiency virus type 1 (HIV-1) acquisition in individuals who are exposed to HIV-1 but do not become infected may provide important direction for the creation of an HIV-1 vaccine. We have employed the simian immunodeficiency virus (SIV)/rhesus monkey model to determine whether monkeys can be repeatedly exposed to a primate lentivirus by a mucosal route and escape infection and whether virus-specific immune correlates of protection from infection can be identified in uninfected monkeys. Five of 18 rhesus monkeys exposed 18 times by intrarectal inoculation to SIVmac251 or SIVsmE660 were resistant to infection, indicating that the exposed/uninfected phenotype can be reproduced in a nonhuman primate AIDS model. However, routine peripheral blood lymphocyte gamma interferon enzyme-linked immunospot (ELISPOT), tetramer, and intracellular cytokine staining assays, as well as cytokine-augmented ELISPOT and peptide-stimulated tetramer assays, failed to define a systemic antigen-specific cellular immune correlate to this protection. Further, local cell-mediated immunity could not be demonstrated by tetramer assays of these protected monkeys, and local humoral immunity was not associated with protection against acquisition of virus in another cohort of mucosally exposed monkeys. Therefore, resistance to mucosal infection in these monkeys may not be mediated by adaptive virus-specific immune mechanisms. Rather, innate immune mechanisms or an intact epithelial barrier may be responsible for protection against mucosal infection in this population of monkeys.     

Roles of target cells and virus-specific cellular immunity in primary simian immunodeficiency virus infection

There is an ongoing debate on whether acute human immunodeficiency virus infection is controlled by target cell limitation or by virus-specific cellular immunity. To resolve this question, we developed a novel mathematical modeling scheme which allows us to incorporate measurements of virus load, target cells, and virus-specific immunity and applied it to a comprehensive data set generated in an experiment involving rhesus macaques infected with simian immunodeficiency virus. Half of the macaques studied were treated during the primary infection period with reagents which block T-cell costimulation and as a result displayed severely impaired virus-specific immune responses. Our results show that early viral replication in normal infection is controlled to a large extent by virus-specific CD8(+) T cells and not by target cell limitation.     

CpG-C immunostimulatory oligodeoxyribonucleotide activation of plasmacytoid dendritic cells in rhesus macaques to augment the activation of IFN-gamma-secreting simian immunodeficiency virus-specific T cells

There are two principle subsets of dendritic cells (DCs); CD11c(+)CD123(-) myeloid DCs (MDCs) and CD11c(-)CD123(+) plasmacytoid DCs (PDCs). DC activation via TNF-TNFRs (e.g., CD40L) and TLRs (e.g., immunostimulatory oligodeoxyribonucleotides (ISS-ODNs)) is crucial for maximal stimulation of innate and adaptive immunity. Macaque DC biology is being studied to improve HIV vaccines using the SIV macaque model. Using lineage (Lin) markers to exclude non-DCs, Lin(-)HLA-DR(+)CD11c(+)CD123(-) MDCs and Lin(-)HLA-DR(+)CD11c(-)CD123(+) PDCs were identified in the blood of uninfected macaques and healthy macaques infected with SIV or simian-human immunodeficiency virus. Overnight culture of DC-enriched Lin-depleted cells increased CD80 and CD86 expression. IL-12 production and CD80/CD86 expression by MDC/PDC mixtures was further enhanced by CD40L and ISS-ODN treatment. A CpG-B ISS-ODN increased CD80/CD86 expression by PDCs, but resulted in little IFN-alpha secretion unless IL-3 was added. In contrast, a CpG-C ISS-ODN and aldrithiol-2-inactivated (AT-2) SIV induced considerable PDC activation and IFN-alpha release without needing exogenous IL-3. The CpG-C ISS-ODN also stimulated IL-12 release (unlike AT-2 SIV) and augmented DC immunostimulatory activity, increasing SIV-specific T cell IFN-gamma production induced by AT-2 SIV-presenting MDC/PDC-enriched mixtures. These data highlight the functional capacities of MDCs and PDCs in naive as well as healthy, infected macaques, revealing a promising CpG-C ISS-ODN-driven DC activation strategy that boosts immune function to augment preventative and therapeutic vaccine efficacy.     

Elicitation of simian immunodeficiency virus-specific cytotoxic T lymphocytes in mucosal compartments of rhesus monkeys by systemic vaccination

Since most human immunodeficiency virus (HIV) infections are initiated following mucosal exposure to the virus, the anatomic containment or abortion of an HIV infection is likely to require vaccine-elicited cellular immune responses in those mucosal sites. Studying vaccine-elicited mucosal immune responses has been problematic because of the difficulties associated with sampling T lymphocytes from those anatomic compartments. In the present study, we demonstrate that mucosal cytotoxic T lymphocytes (CTL) specific for simian immunodeficiency virus (SIV) and simian HIV can be reproducibly sampled from intestinal mucosal tissue of rhesus monkeys obtained under endoscopic guidance. These lymphocytes recognize peptide-major histocompatibility complex class I complexes and express gamma interferon on exposure to peptide antigen. Interestingly, systemic immunization of monkeys with plasmid DNA immunogens followed by live recombinant attenuated poxviruses or adenoviruses with genes deleted elicits high-frequency SIV-specific CTL responses in these mucosal tissues. These studies therefore suggest that systemic delivery of potent HIV immunogens may suffice to elicit substantial mucosal CTL responses.     

Potent simian immunodeficiency virus-specific cellular immune responses in the breast milk of simian immunodeficiency virus-infected, lactating rhesus monkeys

Breast milk transmission of HIV is a leading cause of infant HIV/AIDS in the developing world. Remarkably, only a small minority of breastfeeding infants born to HIV-infected mothers contract HIV via breast milk exposure, raising the possibility that immune factors in the breast milk confer protection to the infants who remain uninfected. To model HIV-specific immunity in breast milk, lactation was pharmacologically induced in Mamu-A*01(+) female rhesus monkeys. The composition of lymphocyte subsets in hormone-induced lactation breast milk was found to be similar to that in natural lactation breast milk. Hormone-induced lactating monkeys were inoculated i.v. with SIVmac251 and CD8(+) T lymphocytes specific for two immunodominant SIV epitopes, Gag p11C and Tat TL8, and SIV viral load were monitored in peripheral blood and breast milk during acute infection. The breast milk viral load was 1-2 logs lower than plasma viral load through peak and set point of viremia. Surprisingly, whereas the kinetics of the SIV-specific cellular immunity in breast milk mirrored that of the blood, the peak magnitude of the SIV-specific CD8(+) T lymphocyte response in breast milk was more than twice as high as the cellular immune response in the blood. Furthermore, the appearance of the SIV-specific CD8(+) T lymphocyte response in breast milk was associated with a reduction in breast milk viral load, and this response remained higher than that in the blood after viral set point. This robust viral-specific cellular immune response in breast milk may contribute to control of breast milk virus replication.     

Durable mucosal simian immunodeficiency virus-specific effector memory T lymphocyte responses elicited by recombinant adenovirus vectors in rhesus monkeys

The induction of potent and durable cellular immune responses in both peripheral and mucosal tissues may be important for the development of effective vaccines against human immunodeficiency virus type 1 and other pathogens. In particular, effector responses at mucosal surfaces may be critical to respond rapidly to incoming mucosal pathogens. Here we report that intramuscular injection of nonreplicating recombinant adenovirus (rAd) vectors into rhesus monkeys induced remarkably durable simian immunodeficiency virus (SIV)-specific T lymphocyte responses that persisted for over 2 years in both peripheral blood and multiple mucosal tissues, including colorectal, duodenal, and vaginal biopsy specimens, as well as bronchoalveolar lavage fluid. In peripheral blood, SIV-specific T lymphocytes underwent the expected phenotypic evolution from effector memory T cells (T(EM)) to central memory T cells (TCM) following vaccination. In contrast, mucosal SIV-specific T lymphocytes exhibited a persistent and durable T(EM) phenotype that did not evolve over time. These data demonstrate that nonreplicating rAd vectors induce durable and widely distributed effector memory mucosal T lymphocyte responses that are phenotypically distinct from peripheral T lymphocyte responses. Vaccine-elicited T(EM) responses at mucosal surfaces may prove critical for affording protection against invading pathogens at the mucosal portals of entry.     

Analysis of TCRalphabeta combinations used by simian immunodeficiency virus-specific CD8+ T cells in rhesus monkeys: implications for CTL immunodominance

Immunodominance is a common feature of Ag-specific CTL responses to infection or vaccines. Understanding the basis of immunodominance is crucial to understanding cellular immunity and viral evasion mechanisms and will provide a rational approach for improving HIV vaccine design. This study was performed comparing CTLs specific for the SIV Gag p11C (dominant) and SIV Pol p68A (subdominant) epitopes that are consistently generated in Mamu-A*01(+) rhesus monkeys exposed to SIV proteins. Additionally, vaccinated monkeys were used to prevent any issues of antigenic variation or dynamic changes in CTL responses by continuous Ag exposure. Analysis of the TCR repertoire revealed the usage of higher numbers of TCR clones by the dominant p11C-specific CTL population. Preferential usage of specific TCRs and the in vitro functional TCR-alpha- and -beta-chain-pairing assay suggests that every peptide/MHC complex may only be recognized by a limited number of unique combinations of alpha- and beta-chain pairs. The wider array of TCR clones used by the dominant p11C-specific CTL population might be explained by the higher probability of generating those specific TCR chain pairs. Our data suggest that Ag-specific naive T cell precursor frequency may be predetermined and that this process dictates immunodominance of SIV-specific CD8(+) T cell responses. These findings will aid in understanding immunodominance and designing new approaches to modulate CTL responses.     

Preservation of functional virus-specific memory CD8+ T lymphocytes in vaccinated, simian human immunodeficiency virus-infected rhesus monkeys

Functional impairment of virus-specific memory CD8(+) T lymphocytes has been associated with clinical disease progression following HIV, SIV, and simian human immunodeficiency virus infection. These lymphocytes have a reduced capacity to produce antiviral cytokines and mediators involved in the lysis of virally infected cells. In the present study, we used polychromatic flow cytometry to assess the frequency and functional capacity of central memory (CD28(+)CD95(+)) and effector memory (CD28(-)CD95(+)) subpopulations of Gag-specific CD8(+) T cells in SIV/simian human immunodeficiency virus-infected rhesus monkeys. The aim of this study was to determine whether Ag-specific, memory CD8(+) T cell function could be preserved in infected monkeys that had been immunized before infection with a vaccine regimen consisting of a plasmid DNA prime followed by a recombinant viral vector boost. We observed that vaccination was associated with the preservation of Gag-specific central memory CD8(+) T cells that were functionally capable of producing IFN-gamma, and effector memory CD8(+) T cells that were capable of producing granzyme B following viral Ag exposure.     

Simian immunodeficiency virus-specific cytotoxic T lymphocytes are present in the AIDS-associated skin rash in rhesus monkeys.

An infiltration of CD8+ lymphocytes in the dermis and epidermis underlies the skin rash that commonly occurs as a primary manifestation of an AIDS virus infection. These cutaneous lymphocytes were characterized in simian immunodeficiency virus of macaques (SIVmac)-infected rhesus monkeys. Skin rash-associated lymphocytes exhibited greater lysis of SIVmac-expressing target cells and a higher cloning efficiency for SIVmac-specific effector T cells than PBL. Moreover, both SIVmac envelope- and gag-specific CTL could be readily cloned from these skin rash-associated lymphocytes. In fact, the skin rash-associated CTL exhibited the same MHC restriction and epitope specificity as those CTL derived from PBL. These studies, therefore, demonstrate that the cutaneous infiltrating CD8+ lymphocytes in SIVmac-infected rhesus monkeys include SIVmac-specific CTL. Thus, whereas virus-specific CTL are likely to represent an important mechanism for controlling AIDS virus infections, they also may play a role in the pathogenesis of the skin lesions that occur after this infection.     

Determination of a T cell receptor of potent CD8+ T cells against simian immunodeficiency virus infection in Burmese rhesus macaques

Cumulative studies on human immunodeficiency virus (HIV)-infected individuals have shown association of major histocompatibility complex class I (MHC-I) polymorphisms with lower viral load and delayed AIDS progression, suggesting that HIV replication can be controlled by potent CD8⁺ T-cell responses. We have previously established an AIDS model of simian immunodeficiency virus (SIV) infection in Burmese rhesus macaques and found a potent CD8⁺ T cell targeting the Mamu-A1*065:01-restricted Gag_241-249 epitope, which is located in a region corresponding to the HIV Gag_240-249 TW10 epitope restricted by a protective MHC-I allele, HLA-B*57. In the present study, we determined a T cell receptor (TCR) of this Gag_241-249 epitope-specific CD8⁺ T cell. cDNA clones encoding TCR-α and TCR-β chains were obtained from a Gag_241-249-specific CD8⁺ T-cell clone. Coexpression of these TCR-α and TCR-β cDNAs resulted in reconstitution of a functional TCR specifically detected by Gag_241-249 epitope-Mamu-A1*065:01 tetramer. Two of three previously-reported CD8⁺ T-cell escape mutations reduced binding affinity of Gag_241-249 peptide to Mamu-A1*065:01 but the remaining one not. This is consistent with the data obtained by molecular modeling of the epitope-MHC-I complex and TCR. These results would contribute to understanding how viral CD8⁺ T-cell escape mutations are selected under structural constraint of viral proteins.     

Killing kinetics of simian immunodeficiency virus-specific CD8+ T cells: implications for HIV vaccine strategies

Both the magnitude and function of vaccine-induced HIV-specific CD8+ CTLs are likely to be important in the outcome of infection. We hypothesized that rapid cytolysis by CTLs may facilitate control of viral challenge. Release kinetics of the cytolytic effector molecules granzyme B and perforin, as well as the expression of the degranulation marker CD107a and IFN-gamma were simultaneously studied in SIV Gag(164-172) KP9-specific CD8+ T cells from Mane-A*10+ pigtail macaques. Macaques were vaccinated with either prime-boost poxvirus vector vaccines or live-attenuated SIV vaccines. Prime-boost vaccination induced Gag-specific CTLs capable of only slow (after 3 h) production of IFN-gamma and with limited (<5%) degranulation and granzyme B release. Vaccination with live-attenuated SIV resulted in a rapid cytolytic profile of SIV-specific CTLs with rapid (<0.5 h) and robust (>50% of tetramer-positive CD8+ T cells) degranulation and granzyme B release. The cytolytic phenotype following live-attenuated SIV vaccinations were similar to that associated with the partial resolution of viremia following SIV(mac251) challenge of prime-boost-vaccinated macaques, albeit with less IFN-gamma expression. High proportions of KP9-specific T cells expressed the costimulatory molecule CD28 when they exhibited a rapid cytolytic phenotype. The delayed cytolytic phenotype exhibited by standard vector-based vaccine-induced CTLs may limit the ability of T cell-based HIV vaccines to rapidly control acute infection following a pathogenic lentiviral exposure.     

Cutting edge: T cells monitor N-myristoylation of the Nef protein in simian immunodeficiency virus-infected monkeys

The use of the host cellular machinery is essential for pathogenic viruses to replicate in host cells. HIV and SIV borrow the host-derived N-myristoyl-transferase and its substrate, myristoyl-CoA, for coupling a saturated C(14) fatty acid (myristic acid) to the N-terminal glycine residue of the Nef protein. This biochemical reaction, referred to as N-myristoylation, assists its targeting to the plasma membrane, thereby supporting the immunosuppressive activity proposed for the Nef protein. In this study, we show that the host immunity is equipped with CTLs capable of sensing N-myristoylation of the Nef protein. A rhesus macaque CD8(+) T cell line was established that specifically recognized N-myristoylated, but not unmodified, peptides of the Nef protein. Furthermore, the population size of N-myristoylated Nef peptide-specific T cells was found to increase significantly in the circulation of SIV-infected monkeys. Thus, these results identify N-myristoylated viral peptides as a novel class of CTL target Ag.     

Functional impairment of simian immunodeficiency virus-specific CD8+ T cells during the chronic phase of infection

In an attempt to determine why high frequencies of circulating virus-specific CD8+ T cells are unable to control human immunodeficiency virus and simian immunodeficiency virus (SIV) replication, we assessed the functional nature of SIV-specific CD8+ lymphocytes. After vaccination and early after infection, nearly all tetramer-staining CD8+ cells produced gamma interferon in response to their specific stimulus. However, by 4 months postinfection with pathogenic SIVmac239, signs of functional impairment in the CD8+ T-cell compartment were detected which might prevent these T cells from efficiently controlling the infection during the chronic phase.     

Plasmacytoid dendritic cells migrate in afferent skin lymph

Conventional dendritic cells enter lymph nodes by migrating from peripheral tissues via the lymphatic route, whereas plasmacytoid dendritic cells (pDC), also called IFN-producing cells (IPC), are described to gain nodes from blood via the high endothelial venules. We demonstrate here that IPC/pDC migrate in the afferent lymph of two large mammals. In sheep, injection of type A CpG oligodinucleotide (ODN) induced lymph cells to produce type I IFN. Furthermore, low-density lymph cells collected at steady state produced type I IFN after stimulation with type A CpG ODN and enveloped viruses. Sheep lymph IPC were found within a minor B(neg)CD11c(neg) subset expressing CD45RB. They presented a plasmacytoid morphology, expressed high levels of TLR-7, TLR-9, and IFN regulatory factor 7 mRNA, induced IFN-gamma production in allogeneic CD4(pos) T cells, and differentiated into dendritic cell-like cells under viral stimulation, thus fulfilling criteria of bona fide pDC. In mini-pig, a CD4(pos)SIRP(pos) subset in afferent lymph cells, corresponding to pDC homologs, produced type I IFN after type A CpG-ODN triggering. Thus, pDC can link innate and acquired immunity by migrating from tissue to draining node via lymph, similarly to conventional dendritic cells.     

Frequency of herpes simplex virus-specific helper T lymphocyte precursors in the lymph node cells of infected mice

We have established a limited dilution assay to estimate the frequency of herpes simplex virus type 1 (HSV)-specific, interleukin 2 (IL 2)-producing helper T lymphocyte precursors (HTL-P). The estimated frequency of such cells in suspensions of local lymph node (LN) cells 5 days after in vivo virus infection was 1:2470 to 1:5800. Frequencies of HTL-P in cells from uninfected mice were below levels of detection of our system and were judged to be below 1:100,000. Removal of Lyt-2+ cells from responder LN cells before culture increases HTL-P frequency twofold to threefold, indicating the likely operation of some form of suppression in unseparated cultures. The demonstration of HSV-specific HTL-P required that cells from virus-primed mice be reexposed in vitro to viral antigen. In addition, clones expanded during a 9-day culture period failed to generate IL 2 unless reexposed to specific viral antigen or cross-reactivate HSV-2. Thus, HSV-specific HTL-P were strictly antigen dependent. No evidence was obtained for antigen-independent subpopulations of HTL-P as occurs with viral-specific cytotoxic T lymphocyte precursors. The clonal progeny of HTL-P were of the Thy-1+ Lyt+2- phenotype. Priming in vivo for the subsequent in vitro detection of HTL-P required that mice be exposed to infectious virus. Thus neither UV-inactivated nor heat-inactivated nor extracted viral glycoproteins could prime for HTL-P detection. The relevance of these findings for the future use of subunit vaccines against HSV is briefly discussed.     

Strongyloides ratti: Transfer of immunity in rats by lymph node cells or serum

Immune mesenteric lymph node cells (IMLNC) and hyperimmune serum, alone or in combination, were transferred to recipient, syngeneic Lewis rats. On the day of cell transfer, the recipients, along with appropriate controls, were infected with 1,000Strongyloides ratti larvae. Twelve days later, the number of adult worms in the anterior small intestine was counted and the number ofS. ratti eggs in the terminal uterus of representative female worms was determined. Using these methods, we showed that worm fecundity was reduced in the IMLNC recipients, the hyperimmune serum recipients, and the recipients of both cells and serum as compared with regular controls (averages of 2.45, 2.93, and 2.98 eggs, respectively, vs. 4.73 eggs). The number of adult worms recovered from the IMLNC recipients was not reduced (474.6 worms). However, significantly fewer worms were recovered from the recipients of IMLNC-hyperimmune serum and the recipients of hyperimmune serum only as compared with regular controls (184.8 and 171.5 worms, respectively, vs. 567.2 worms). A working hypothesis, involving an interaction of serum and cellular components, is postulated as the mechanism for this protective response.     

Increased macrophage infection upon subcutaneous inoculation of rhesus macaques with simian immunodeficiency virus-loaded dendritic cells or T cells but not with cell-free virus

Information on the establishment of immunodeficiency virus infection through transmission of infected cells is sparse. Dendritic cells (DCs) and T cells may be central to the onset and subsequent spread of infection following mucosal exposure. To directly investigate the consequences of virus being introduced by DCs or T cells, we reinjected ex vivo simian immunodeficiency virus (SIV)-loaded autologous immature DCs and T cells subcutaneously (s.c.) into healthy macaques. s.c. injection of cell-bound virus was used to mirror what may happen if virus-loaded cells pass through an epithelium or perhaps DCs and T cells that immediately entrap cell-free virus, having just crossed an epithelial barrier. Virus load in the plasma was monitored along with combined in situ hybridization and immunohistochemistry to identify the cells replicating virus in the lymphoid tissues. Both DCs and T cells transmitted infection after being pulsed with either wild-type or nef-defective (delta nef) SIVmac239. As seen in animals infected intravenously, replication of delta nef was attenuated compared to that of wild-type virus when introduced in either cell-bound form. Upon examination of the draining lymph nodes (LNs) during the first days of infection, virus-producing CD4(+) T cells predominated in control animals that received s.c. cell-free virus. In dramatic contrast, both SIV-positive macrophages and T cells were detected in the LNs of monkeys infected with cell-associated SIV. Therefore, although both cell-free and cell-associated viruses are infectious, the initial cells amplifying the virus differ. This may have important implications for the subsequent dissemination of infection and/or induction of antiretroviral immunity.     

Emergence and kinetics of simian immunodeficiency virus-specific CD8(+) T cells in the intestines of macaques during primary infection

In this report, three Mamu-A*01(+) rhesus macaques were examined to compare the emergence of simian immunodeficiency virus (SIV)-specific CD8(+) T cells in the intestines and blood in early SIV infection using a major histocompatibility complex class I tetramer complexed with the Gag(181-189) peptide. Fourteen days after intravenous inoculation with SIVmac251, large numbers of SIV Gag(181-189)-specific CD8(+) T cells were detected in the intestinal mucosa (3.1 to 11.5% of CD3(+) CD8(+) lymphocytes) as well as in the blood (3.1 to 13.4%) of all three macaques. By 21 days postinoculation, levels of tetramer-binding cells had dropped in both the intestines and blood. At day 63, however, levels of SIV Gag(181-189)-specific CD8(+) T cells in the intestines had rebounded in all three macaques to levels that were higher (8.6 to 18.7%) than those at day 21. In contrast, percentages of tetramer-binding cells in the peripheral blood remained comparatively stable (2.5 to 4.5%) at this time point. In summary, SIV Gag(181-189)-specific CD8(+) T cells appeared in both the intestinal mucosa and peripheral blood at a comparable rate and magnitude in primary SIV infection. Given that the intestine is a major site of early viral replication as well as the site where most of the total body lymphocyte pool resides, these data indicate that it is also an early and important site of development of antiviral immune responses.     

Failure or success in the restoration of virus-specific cytotoxic T lymphocyte response defects by dendritic cells

C57BL/6 (B6, H-2b) mice are CTL responders to both Sendai virus and Moloney leukemia virus. In the former response the H-2Kb class I MHC molecule is used as CTL restriction element, in the latter response the H-2Db molecule. B6 dendritic cells (DC) are superior in the presentation of Sendai virus Ag to CTL in comparison with B6 normal spleen cells. Con A blasts have even less capacity to present viral Ag than NSC, and LPS blasts show an intermediate capacity to present viral Ag. H-2Kb mutant bm1 mice do not generate a CTL response to Sendai virus, but respond to Moloney leukemia virus, as demonstrated by undetectable CTL precursors to Sendai virus and a normal CTL precursor frequency to Moloney virus. Compared to B6 mice, other H-2Kb mutant mice show decreased Sendai virus-specific CTL precursor frequencies in a hierarchy reflecting the response in bulk culture. The Sendai virus-specific CTL response defect of bm1 mice was not restored by highly potent Sendai virus-infected DC as APC for in vivo priming and/or in vitro restimulation. In mirror image to H-2Kb mutant bm1 mice, H-2Db mutant bm14 mice do not generate a CTL response to Moloney virus, but respond normally to Sendai virus. This specific CTL response defect was restored by syngeneic Moloney virus-infected DC for in vitro restimulation. This response was Kb restricted indicating that the Dbm14 molecule remained largely defective and that a dormant Kb repertoire was aroused after optimal Ag presentation by DC. In conclusion, DC very effectively present viral Ag to CTL. However, their capacity to restore MHC class I determined specific CTL response defects probably requires at least some ability of a particular MHC class I/virus combination to associate and thus form an immunogenic complex.