Herpes simplex virus type 1-specific cytotoxic T-lymphocyte arming occurs within lymph nodes draining the site of cutaneous infection

Various studies have shown that major histocompatibility complex class I-restricted cytotoxic T lymphocytes (CTL) can be isolated from lymph nodes draining sites of cutaneous infection with herpes simplex virus type 1 (HSV-1). Invariably, detection of this cytolytic activity appeared to require some level of in vitro culture of the isolated lymph node cells, usually for 3 days, in the absence of exogenous viral antigen. This in vitro "resting" period was thought to represent the phase during which committed CD8(+) T cells become "armed" killers after leaving the lymph nodes and prior to their entry into infected tissue as effector CTL. In this study we reexamined the issue of CTL appearance in the HSV-1 immune response and found that cytolytic activity can be isolated directly from draining lymph nodes, although at levels considerably below those found after in vitro culture. By using T-cell receptor elements that represent effective markers for class I-restricted T cells specific for an immunodominant glycoprotein B (gB) determinant from HSV-1, we show that the increase in cytotoxicity apparent after in vitro culture closely mirrors the expansion of gB-specific CTL during the same period. Taken together, our results suggest that HSV-1-specific CTL priming does not appear to require any level of cytolytic machinery arming outside the lymph node compartment despite the absence of any detectable infection within that site.     

Expression of intracellular IFN-gamma in HSV-1-specific CD8+ T cells identifies distinct responding subpopulations during the primary response to infection

Cutaneous infection in the footpads of C57BL/6 mice with HSV-1 results in an accumulation of activated (CD44high CD25+) CD8+ T cells within the draining popliteal lymph node (PLN). These studies were undertaken to evaluate the frequency and phenotype of the CD8+ T cell population within the PLN, recognizing the single immunodominant HSV-1 epitope derived from the viral envelope glycoprotein, glycoprotein B (gB), using an intracellular IFN-gamma-staining assay. It revealed that approximately 6% of the CD8+ T cells were specific for the gB epitope. Phenotypic analysis of the IFN-gamma-producing gB-specific CD8+ T cells generated in the PLN during the course of the acute infection expressed the CD44high CD25+ phenotype on days 3-5 postinfection. Surprisingly, IFN-gamma-producing CD8+ T cells expressed the CD44high CD25- phenotype on days 5-8 postinfection, in contrast to expectations for a CD8+ effector T cell. IFN-gamma-producing CD25- CD8+ T cells were detected in the PLN on day 21 postinfection, long after infectious virus had been cleared. Throughout the response, the spleen was found to be the major reservoir of gB-specific CD8+ T cells, even during the peak of the response. In contrast to the gB-specific CD8+ T cell population within the PLN, the entire gB-specific CD8+ T cell population within the spleen was CD25-. Collectively, these results suggest the generation of subpopulations of virus-specific CD8+ T cells, distinguished by the expression of CD25, during the acute phase of the primary response to a localized viral infection.     

Phenotypic characteristics associated with the acquisition of HSV-specific CD8 T-lymphocyte-mediated cytolytic function in vitro.

Class I MHC-restricted, HSV-1-specific CD8(+) cytolytic T lymphocyte (CTL) function is rarely detected in lymphocytes isolated directly from the lymph node draining the site of infection. However, culture in vitro for 24 to 72 h in the absence of exogenous antigen results in the development of easily detectable levels of HSV-1-specific CTL effectors. The inability to detect virus-specific CTL in HSV-1-infected mice is not well understood. However, since the in vitro culture of HSV-1-immune lymphocytes results in the transition to CTL function, studies of the changes occurring to the CD8(+) T cell subpopulation may provide important insights into the development of virus-specific CTL. Therefore, the phenotypic changes taking place in the CD8(+) population of T cells from draining popliteal lymph nodes of HSV-1-infected C57BL/6 (B6) mice were investigated, focusing on changes in the expression of cell surface markers associated with T lymphocyte activation. The results demonstrate an increase in the percentage of CD8(+) T cells expressing the activation markers CD44 and CD25 in parallel with the acquisition of HSV-specific CTL effector function. Cytolytic function was found exclusively within the CD8(+) CD44(hi) CD25(hi) fraction of cells in culture, but, surprisingly, was not detectable in CD8(+) CD44(hi) CD25(lo) T cells. This suggested that the acquisition of high levels of the high-affinity IL-2 receptor was closely linked to cytolytic function and may define an important developmental stage in the transition from noncytolytic to cytolytic effector cell. In support of this, CD8(+) CD25(hi) T cells isolated from the regional lymph node exhibited direct ex vivo cytolytic function, indicating that cytolytic effector cells were present in the lymph node, but must emigrate rapidly after attaining this level of differentiation.     

Activation, differentiation, and migration of naive virus-specific CD8+ T cells during pulmonary influenza virus infection

The low precursor frequency of individual virus-specific CD8(+) T cells in a naive host makes the early events of CD8(+) T cell activation, proliferation, and differentiation in response to viral infection a challenge to identify. We have therefore examined the response of naive CD8(+) T cells to pulmonary influenza virus infection with a murine adoptive transfer model using hemagglutinin-specific TCR transgenic CD8(+) T cells. Initial activation of CD8(+) T cells occurs during the first 3 days postinfection exclusively within the draining lymph nodes. Acquisition of CTL effector functions, including effector cytokine and granule-associated protease expression, occurs in the draining lymph nodes and differentially correlates with cell division. Division of activated CD8(+) T cells within the draining lymph nodes occurs in an asynchronous manner between days 3 and 4 postinfection. Despite the presence of Ag for several days within the draining lymph nodes, dividing T cells do not appear to maintain contact with residual Ag. After multiple cell divisions, CD8(+) T cells exit the draining lymph nodes and migrate to the infected lung. Activated CD8(+) T cells also disseminate throughout lymphoid tissue including the spleen and distal lymph nodes following their emigration from draining lymph nodes. These results demonstrate an important role for draining lymph nodes in orchestrating T cell responses during a local infection of a discrete organ to generate effector CD8(+) T cells capable of responding to infection and seeding peripheral lymphoid tissues.     

Cross presentation of antigen on MHC class II via the draining lymph node after corneal transplantation in mice

We investigated Ag trafficking from the cornea and T effector cell activation in secondary lymphoid tissue after corneal transplantation. In preliminary experiments, the central cornea was shown to contain a population of CD45(+), CD11b(+), CD11c- cells, with a few MHC class II(+) cells, and F4/80(+) cells. However, MHC class II(+) passenger leukocytes in donor cornea after allografting did not traffic to the draining lymph node. Instead, Ag (plasmid) delivered to the eye via the donor cornea during allograft was detected in host CD11c(+) and F4/80(+) APC in the draining lymph nodes and spleen. The earliest detection of APC-associated Ag was at 6 h in the draining lymph node and 24 h in the spleen. After 48 h Ag was not detected in the draining lymph node but was still present in the spleen. Ag applied to the donor corneal epithelium before allografting induced Ag-specific T cell activation and expansion in the draining lymph node with a peak response at 4-6 days, indicating that cross-presentation of Ag had occurred. We conclude therefore, that Ag is transported from the donor cornea within host APC and that this event occurs within hours after grafting. Ag is cross-presented to host CD4(+) T cells on MHC class II and leads to the activation of Ag-specific effector T cells and clonal expansion in the draining lymph node.     

Induction of rapid T cell activation, division, and recirculation by intratracheal injection of dendritic cells in a TCR transgenic model

Dendritic cells (DCs) are thought to be responsible for sensitization to inhaled Ag and induction of adaptive immunity in the lung. The characteristics of T cell activation in the lung were studied after transfer of Ag-pulsed bone marrow-derived DCs into the airways of naive mice. Cell division of Ag-specific T cells in vivo was followed in a carboxyfluorescein diacetate succinimidyl ester-labeled cohort of naive moth cytochrome c-reactive TCR transgenic T cells. Our adoptive transfer system was such that transferred DCs were the only cells expressing the MHC molecule required for presentation of cytochrome c to transgenic T cells. Ag-specific T cell activation and proliferation occurred rapidly in the draining lymph nodes of the lung, but not in nondraining lymph nodes or spleen. No bystander activation of non-Ag-specific T cells was induced. Division of Ag-specific T cells was accompanied by transient expression of CD69, while up-regulation of CD44 increased with each cell division. Divided cells had recirculated to nondraining lymph nodes and spleen by day 4 of the response. In vitro restimulation with specific Ag revealed that T cells were primed to proliferate more strongly and to produce higher amounts of cytokines per cell. These data are consistent with the notion that DCs in the lung are extremely efficient in selecting Ag-reactive T cells from a diverse repertoire. The response is initially localized in the mediastinal lymph nodes, but subsequently spreads systemically. This system should allow us to study the early events leading to sensitization to inhaled Ag.     

Cutting edge: conventional CD8 alpha+ dendritic cells are preferentially involved in CTL priming after footpad infection with herpes simplex virus-1

CTL play a major role in immunity to HSV type 1, but little is known about the priming process. In this study, we have examined the class I-restricted presentation of an immunodominant determinant from HSV-1 glycoprotein B after footpad infection. We have found that the only cell types capable of presenting this determinant in draining popliteal lymph nodes within the first 3 days after infection are the CD11c(+)CD8alpha(+)CD45RA(-) dendritic cells. Given that such class I-restricted presentation is essential for CTL priming, this implies that these conventional CD8alpha(+) dendritic cells are the key subset involved in CTL immunity to this virus.     

The Initial Draining Lymph Node Primes the Bulk of the CD8 T Cell Response and Influences Memory T Cell Trafficking after a Systemic Viral Infection

Lymphocytic choriomeningitis virus (LCMV) causes a systemic infection in mice with virus replication occurring in both peripheral tissues and secondary lymphoid organs. Because of the rapid systemic dissemination of the virus, the secondary lymphoid organs responsible for the induction of the LCMV-specific CD8 T cell response are poorly defined. We show that the mediastinal lymph node (MedLN) serves as the primary draining lymph node following LCMV infection. In addition, we demonstrate that the MedLN is responsible for priming the majority of the virus-specific CD8 T cell response. Following resolution of the acute infection, the draining MedLN exhibits characteristics of a reactive lymph node including an increased presence of germinal center B cells and increased cellularity for up to 60 days post-infection. Furthermore, the reactive MedLN harbors an increased frequency of CD62L⁻ effector memory CD8 T cells as compared to the non-draining lymph nodes. The accumulation of LCMV-specific CD62L⁻ memory CD8 T cells in the MedLN is independent of residual antigen and is not a unique feature of the MedLN as footpad infection with LCMV leads to a similar increase of virus-specific CD62L⁻ effector memory CD8 T cells in the draining popliteal lymph node. Our results indicate that CD62L⁻ effector memory CD8 T cells are granted preferential access into the draining lymph nodes for an extended time following resolution of an infection.     

Modeling of influenza-specific CD8+ T cells during the primary response indicates that the spleen is a major source of effectors

The biological parameters that determine the distribution of virus-specific CD8(+) T cells during influenza infection are not all directly measurable by experimental techniques but can be inferred through mathematical modeling. Mechanistic and semimechanistic ordinary differential equations were developed to describe the expansion, trafficking, and disappearance of activated virus-specific CD8(+) T cells in lymph nodes, spleens, and lungs of mice during primary influenza A infection. An intensive sampling of virus-specific CD8(+) T cells from these three compartments was used to inform the models. Rigorous statistical fitting of the models to the experimental data allowed estimation of important biological parameters. Although the draining lymph node is the first tissue in which Ag-specific CD8(+) T cells are detected, it was found that the spleen contributes the greatest number of effector CD8(+) T cells to the lung, with rates of expansion and migration that exceeded those of the draining lymph node. In addition, models that were based on the number and kinetics of professional APCs fit the data better than those based on viral load, suggesting that the immune response is limited by Ag presentation rather than the amount of virus. Modeling also suggests that loss of effector T cells from the lung is significant and time dependent, increasing toward the end of the acute response. Together, these efforts provide a better understanding of the primary CD8(+) T cell response to influenza infection, changing the view that the spleen plays a minor role in the primary immune response.     

CXCL9 and CXCL10 expression are critical for control of genital herpes simplex virus type 2 infection through mobilization of HSV-specific CTL and NK cells to the nervous system

CXCL9 and CXCL10 mediate the recruitment of T lymphocytes and NK cells known to be important in viral surveillance. The relevance of CXCL10 in comparison to CXCL9 in response to genital HSV-2 infection was determined using mice deficient in CXCL9 (CXCL9-/-) and deficient in CXCL10 (CXCL10-/-) along with wild-type (WT) C57BL/6 mice. An increased sensitivity to infection was found in CXCL10-/- mice in comparison to CXCL9-/- or WT mice as determined by detection of HSV-2 in the CNS at day 3 postinfection. However, by day 7 postinfection both CXCL9-/- and CXCL10-/- mice possessed significantly higher viral titers in the CNS in comparison to WT mice consistent with mortality (18-35%) of these mice within the first 7 days after infection. Even though CXCL9-/- and CXCL10-/- mice expressed elevated levels of CCL2, CCL3, CCL5, and CXCL1 in the spinal cord in comparison to WT mice, there was a reduction in NK cell and virus-specific CD8+ T cell mobilization to this tissue, suggesting CXCL9 and CXCL10 are critical for recruitment of these effector cells to the spinal cord following genital HSV-2 infection. Moreover, leukocytes from the spinal cord but not from draining lymph nodes or spleens of infected CXCL9-/- or CXCL10-/- mice displayed reduced CTL activity in comparison to effector cells from WT mice. Thus, the absence of CXCL9 or CXCL10 expression significantly alters the ability of the host to control genital HSV-2 infection through the mobilization of effector cells to sites of infection.     

The suppressor T cell induced by syngeneic splenic cell antigen down-regulates hapten-specific cytotoxic T cells by elaborating a factor inhibitory for IL2-dependent cell replication

An in vitro study has been made of the mechanism by which a suppressor T cell, that is induced in lymph nodes by a syngeneic splenic cell antigen, prevents generation of cytotoxic T cells specific for hapten-altered self antigens. When popliteal lymph node cells exposed in vivo to syngeneic splenic cells were immunized in vitro with heat-treated syngeneic TNP-coupled thymocytes and excess helper factors, the Ts remained inactive. In this condition the exposed popliteal lymph node cells routinely demonstrated approximately twice the CTL response developed by lymph node cells from normal mice. Nevertheless, when triggered in vitro by splenic antigen on either X-irradiated B or T cells, the exposed but not the normal lymph node cells exhibited reduced hapten-altered self-specific CTL responses. Furthermore, T cells within spleen cell-exposed popliteal lymph node cell populations when reexposed to splenic T cells made a factor that was found to be suppressive of CTL generation by normal lymph node cells in vitro. The nondialyzable T-cell suppressor factor (TsF) did not appear to act on lymph node precursor CTLs, nor on helper T cells but instead acted at the level of utilization of helper factors in the development of CTLs. In an examination of the effect of TsF on cellular replication, TsF was found to be nontoxic for CTLL-20, an IL-2-dependent T cell, and it did not hinder the uptake of IL-2 by receptor blockade of this cell. Nevertheless, the replication of CTLL-20 that is IL-2 driven was diminished in the presence of TsF. Similarly, TsF was found to be inhibitory for T-cell proliferation stimulated by mitogen but had no effect on a B myeloma cell proliferative response. Thus, TsF appears to act as an inhibitor of a T cell's capability to replicate despite the presence of the stimulus for replication, namely, IL-2.     

CD8+ T cell-dependent elimination of dendritic cells in vivo limits the induction of antitumor immunity

The fate of dendritic cells (DC) after they have initiated a T cell immune response is still undefined. We have monitored the migration of DC labeled with a fluorescent tracer and injected s.c. into naive mice or into mice with an ongoing immune response. DC not loaded with Ag were detected in the draining lymph node in excess of 7 days after injection with maximum numbers detectable approximately 40 h after transfer. In contrast, DC that had been loaded with an MHC class I-binding peptide disappeared from the lymph node with kinetics that parallel the known kinetics of activation of CD8+ T cells to effector function. In the presence of high numbers of specific CTL precursors, as in TCR transgenic mice, DC numbers were significantly decreased by 72 h after injection. The rate of DC disappearance was extremely rapid and efficient in recently immunized mice and was slower in "memory" mice in which memory CD8+ cells needed to reacquire effector function before mediating DC elimination. We also show that CTL-mediated clearance of Ag-loaded DC has a notable effect on immune responses in vivo. Ag-specific CD8+ T cells failed to divide in response to Ag presented on a DC if the DC were targets of a pre-existing CTL response. The induction of antitumor immunity by tumor Ag-loaded DC was also impaired. Therefore, CTL-mediated clearance of Ag-loaded DC may serve as a negative feedback mechanism to limit the activity of DC within the lymph node.     

A novel functional CTL avidity/activity compartmentalization to the site of mucosal immunization contributes to protection of macaques against simian/human immunodeficiency viral depletion of mucosal CD4+ T cells

The presence of high-avidity CTLs in the right compartment can greatly affect clearance of a virus infection (for example, AIDS viral infection of and dissemination from mucosa). Comparing mucosal vs systemic immunization, we observed a novel compartmentalization of CTL avidity and proportion of functionally active Ag-specific CD8(+) T cells to tissues proximal to sites of immunization. Whereas both s.c. and intrarectal routes of immunization induced tetramer(+) cells in the spleen and gut, the mucosal vaccine induced a higher percentage of functioning IFN-gamma(+) Ag-specific CD8(+) T cells in the gut mucosa in mice. Translating to the CD8(+) CTL avidity distribution in rhesus macaques, intrarectal vaccination induced more high-avidity mucosal CTL than s.c. vaccination and protection of mucosal CD4(+) T cells from AIDS viral depletion, whereas systemic immunization induced higher avidity IFN-gamma-secreting cells in the draining lymph nodes but no protection of mucosal CD4(+) T cells, after mucosal challenge with pathogenic simian/human immunodeficiency virus. Mucosal CD4(+) T cell loss is an early critical step in AIDS pathogenesis. The preservation of CD4(+) T cells in colonic lamina propria and the reduction of virus in the intestine correlated better with high-avidity mucosal CTL induced by the mucosal AIDS vaccine. This preferential localization of high-avidity CTL may explain previous differences in vaccination results and may guide future vaccination strategy.     

Characterization of lymphocyte responsiveness in early experimental syphilis. I. In vitro response to mitogens and Treponema pallidum antigens

Lymphoid cells from spleens and lymph nodes of rabbits infected with T. pallidum respond by proliferation to concanavalin A (Con A) and T. pallidum antigens. Spleen cell responsiveness to treponemal antigens appears 6 days after infection, is 100 to 600 fold higher than the response of uninfected control rabbits, and is maintained throughout the 31-day observation period. Specifically responding cells in the inguinal and popliteal lymph nodes of infected animals are demonstrable on day 10, and the magnitude of the response increases throughout the observation period. Specific responsiveness to T. pallidum antigens in vitro is enhanced in purified T cell populations and is abolished by treatment with goat anti-rabbit thymocyte serum and complement. The response of spleen and lymph node cells to Con A is unaffected during syphilitic infection. These results are consistent with a role for T cell-mediated specific immunity to treponemal antigens early after infection and do not support a hypothesis of depressed cellular immunity during syphilitic infection.     

The antimetastatic function of concomitant antitumor immunity. II. Evidence that the generation of Ly-1+2+ effector T cells temporarily causes the destruction of already disseminated tumor cells

Progressive growth of the P815 mastocytoma in an immunocompetent host evokes the generation of an antitumor immune response that can be measured in terms of the production of cytolytic Ly-1+2+ T cells in the draining lymph node and spleen. This immunity, designated concomitant immunity, is present on day 6 of tumor growth, peaks on day 9, and decays progressively thereafter. It fails to develop in mice made T cell deficient by thymectomy and lethal whole-body gamma-radiation, and reconstituted with syngeneic bone marrow cells (TXB mice). Employment of a mouse survival assay, capable of enumerating metastatic P815 cells in cell suspensions, showed that the P815 tumor metastasizes to the draining lymph node and spleen at the same rate in normal and TXB mice for the first 6 days of growth of an intradermal P815 tumor. By day 6 of tumor growth there were approximately 10(3) P815 cells in the draining lymph node in both types of mice. However, during the generation of concomitant immunity between days 6 and 9, the number of metastatic P815 cells in the draining lymph nodes and spleens of normal tumor-bearing mice declined by nearly 90%. After day 12, however, the number of tumor cells in the nodes and spleens increased concordantly with the decay of concomitant immunity. These findings, together with the demonstration that T cell-deficient mice failed to restrain the number of metastatic P815 cells in the draining lymph node and spleen, suggest that concomitant immunity is an important defense mechanism against the development of systemic disease. Additional evidence consistent with this interpretation was provided by studies which showed that adoptive immunization with spleen cells from concomitant immune donors significantly prolonged the median survival time of TXB tumor-bearing mice by destroying a substantial proportion of P815 tumor cells already seeded in the draining lymph node. Adoptive immunization also delayed the appearance of metastatic tumor cells in the spleen.     

Naive, effector, and memory CD8 T cells in protection against pulmonary influenza virus infection: homing properties rather than initial frequencies are crucial.

The goal of adoptive immunotherapy is to target a high number of persisting effector cells to the site of a virus infection or tumor. In this study, we compared the protective value of hemagglutinin peptide-specific CD8 T cells generated from the clone-4 TCR-transgenic mice, defined by different stages of their differentiation, against lethal pulmonary influenza infection. We show that the adoptive transfer of high numbers of Ag-specific unprimed, naive CD8 T cells failed to clear the pulmonary virus titer and to promote host survival. The same numbers of in vitro generated primary Ag-specific Tc1 effector cells, producing high amounts of IFN-gamma, or resting Tc1 memory cells, generated from these effectors, were protective. Highly activated CD62Llow Tc1 effectors accumulated in the lung with rapid kinetics and most efficiently reduced the pulmonary viral titer early during infection. The resting CD62Lhigh naive and memory populations first increased in cell numbers in the draining lymph nodes. Subsequently, memory cells accumulated more rapidly and to a greater extent in the lung lavage as compared with naive cells. Thus, effector cells are most effective against a localized virus infection, which correlates with their ability to rapidly distribute at the infected tissue site. The finding that similar numbers of naive Ag-specific CD8 T cells are not protective supports the view that qualitative differences between the two resting populations, the naive and the memory population, may play a major role in their protective value against disease.     

Functional responses and costimulator dependence of memory CD4+ T cells

To examine the functional characteristics of memory CD4+ T cells, we used an adoptive transfer system to generate a stable population of Ag-specific memory cells in vivo and compared their responses to Ag with those of a similar population of Ag-specific naive cells. Memory cells localized to the spleen and lymph nodes of mice and exhibited extremely rapid recall responses to Ag in vivo, leaving the spleen within 3-5 days of Ag encounter. Unlike their naive counterparts, memory cells produced effector cytokines (IFN-gamma, IL-4, IL-5) within 12-24 h of Ag exposure and did not require multiple cycles of cell division to do so. Memory cells proliferated at lower Ag concentrations than did naive cells, were less dependent on costimulation by B7 molecules, and independent of costimulation by CD40. Furthermore, effector cytokine production by memory cells also occurred in the absence of either B7 or CD40 costimulation. Lastly, memory cells were resistant to tolerance induction. Together, these findings suggest that the threshold for activation of memory CD4+ cells is lower than that of naive cells. This would permit memory cells to rapidly express their effector functions in vivo earlier in the course of a secondary immune response, when the levels of Ag and the availability of costimulation may be relatively low.     

MAdCAM-1 expressing sacral lymph node in the lymphotoxin beta-deficient mouse provides a site for immune generation following vaginal herpes simplex virus-2 infection

The members of the lymphotoxin (LT) family of molecules play a critical role in lymphoid organogenesis. Whereas LT alpha-deficient mice lack all lymph nodes and Peyer's patches, mice deficient in LT beta retain mesenteric lymph nodes and cervical lymph nodes, suggesting that an LT beta-independent pathway exists for the generation of mucosal lymph nodes. In this study, we describe the presence of a lymph node in LT beta-deficient mice responsible for draining the genital mucosa. In the majority of LT beta-deficient mice, a lymph node was found near the iliac artery, slightly misplaced from the site of the sacral lymph node in wild-type mice. The sacral lymph node of the LT beta-deficient mice, as well as that of the wild-type mice, expressed the mucosal addressin cell adhesion molecule-1 similar to the mesenteric lymph node. Following intravaginal infection with HSV type 2, activated dendritic cells capable of stimulating a Th1 response were found in this sacral lymph node. Furthermore, normal HSV-2-specific IgG responses were generated in the LT beta-deficient mice following intravaginal HSV-2 infection even in the absence of the spleen. Therefore, an LT beta-independent pathway exists for the development of a lymph node associated with the genital mucosa, and such a lymph node serves to generate potent immune responses against viral challenge.     

An hour after immunization peritoneal B-1 cells are activated to migrate to lymphoid organs where within 1 day they produce IgM antibodies that initiate elicitation of contact sensitivity

Elicitation of contact sensitivity (CS), a classic example of T cell-mediated immunity, requires Ag-specific IgM Abs to trigger an initiation process. This early process leads to local recruitment of CS-effector T cells after secondary Ag challenge. These Abs are produced by the B-1 subset of B cells within 1 day after primary skin immunization. In this study we report the surprising observation that B-1 cells in the peritoneal cavity are activated as early as 1 h after naive mice are painted with a contact-sensitizing Ag on the skin of the trunk and feet to begin the initiation of CS. B-1 cells in the spleen and draining lymph nodes produce the initiating Abs by 1 day after immunization, when we found increased numbers of Ag-specific IgM Ab-producing cells in these tissues by ELISPOT assay. Importantly, we show that contact-activated peritoneal B-1 cells migrate to these lymphoid tissues and then differentiate into Ag-specific IgM Ab-producing cells, resulting in specific CS-initiating IgM Abs in the serum by 1 day. Furthermore, pertussis toxin, which is known to inhibit signaling via G protein-coupled chemokines, inhibited the migration of contact-activated peritoneal B-1 cells to the lymphoid tissues, probably due to BLR-1 (Burkitt lymphoma receptor-1). These findings indicate that within 1 h after contact skin immunization, B-1 cells in the peritoneal cavity are activated to migrate to the lymphoid tissues by chemokine-dependent mechanisms to produce serum Ag-specific IgM Abs within 1 day after immunization, leading to local recruitment of CS-effector T cells.     

The early expression of glycoprotein B from herpes simplex virus can be detected by antigen-specific CD8+ T cells

The immune response to cutaneous herpes simplex virus type 1 (HSV-1) infection begins with remarkable rapidity. Activation of specific cytotoxic T lymphocytes (CTL) begins within hours of infection, even though the response within the draining lymph nodes peaks nearly 5 days later. HSV gene products are classified into three main groups, alpha, beta, and gamma, based on their kinetics and requirements for expression. In C57BL/6 mice, the immunodominant epitope from HSV is derived from glycoprotein B (gB(498-505)). While gB is considered a gamma or "late" gene product, previous reports have indicated that some level of gene expression may occur soon after infection. Using brefeldin A as a specific inhibitor of viral antigen presentation to major histocompatibility complex class I-restricted CTL, we have formally addressed the timing of gB peptide expression in an immunologically relevant manner following infection. Presentation of gB peptide detected by T-cell activation was first observed within 2 h of infection. Comparison with another viral epitope expressed early during infection, HSV-1 ribonucleotide reductase, demonstrated that gB is presented with the same kinetics as this classical early-gene product. Moreover, this rapidity of gB expression was further illustrated via rapid priming of na?ve transgenic CD8(+) T cells in vivo after HSV-1 infection of mice. These results establish that gB is expressed rapidly following HSV-1 infection, at levels capable of effectively stimulating CD8(+) T cells.