Antigen modulation of the immune response. VI. Rate of large memory cell appearance in lymph nodes and thoracic duct lymph

We have studied the rate of appearance of memory B-cell subpopulations in the antigen draining lymph nodes and thoracic duct lymph of rats using 1g velocity sedimentation and adoptive transfer. Five days after immunization 100% of the memory response was attributable to large cells. By Days 7, 14, 28, and 77 after priming the large cells contribution to the memory population dropped to 86, 35, 15, and 10% respectively. At the same time the small cell contribution rose from 20% on Day 14 to 46% on Days 28 and 77. The same results were obtained with thoracic duct lymphocytes with the large cells contributing 53% of the response on Day 7 and 20% on Day 150. Appropriate controls were included to show that differential suppression was not responsible for these results. Furthermore, when purified large memory cells were passaged through intermediate hosts for 7 to 11 days, between 76 and 81% of the large cells matured into medium or small lymphocytes. These data show that the earliest memory cells formed after antigen encounter are the blast-like large lymphocytes and that these evolve, through a series of antigen-independent events, first into medium and then small lymphocytes. A model of memory cell development incorporating these results and the results of others is presented.     

Distribution of primed T cells and antigen-loaded antigen presenting cells following intranasal immunization in mice

Priming of T cells is a key event in vaccination, since it bears a decisive influence on the type and magnitude of the immune response. T-cell priming after mucosal immunization via the nasal route was studied by investigating the distribution of antigen-loaded antigen presenting cells (APCs) and primed antigen-specific T cells. Nasal immunization studies were conducted using the model protein antigen ovalbumin (OVA) plus CpG oligodeoxynucleotide adjuvant. Trafficking of antigen-specific primed T cells was analyzed in vivo after adoptive transfer of OVA-specific transgenic T cells in the presence or absence of fingolimod, a drug that causes lymphocytes sequestration within lymph nodes. Antigen-loaded APCs were observed in mediastinal lymph nodes, draining the respiratory tract, but not in distal lymph nodes. Antigen-specific proliferating T cells were first observed within draining lymph nodes, and later in distal iliac and mesenteric lymph nodes and in the spleen. The presence at distal sites was due to migration of locally primed T cells as shown by fingolimod treatment that caused a drastic reduction of proliferated T cells in non-draining lymph nodes and an accumulation of extensively divided T cells within draining lymph nodes. Homing of nasally primed T cells in distal iliac lymph nodes was CD62L-dependent, while entry into mesenteric lymph nodes depended on both CD62L and α4β7, as shown by in vivo antibody-mediated inhibition of T-cell trafficking. These data, elucidating the trafficking of antigen-specific primed T cells to non-draining peripheral and mucosa-associated lymph nodes following nasal immunization, provide relevant insights for the design of vaccination strategies based on mucosal priming.     

Progression of armed CTL from draining lymph node to spleen shortly after localized infection with herpes simplex virus 1.

We have examined the generation of CTL immunity immediately after localized footpad infection with herpes simplex virus 1 (HSV-1) using three coordinated in vivo T cell tracking methodologies. Tetrameric MHC class I containing the immunodominant peptide from HSV-1 glycoprotein B (gB) showed that after infection the proportion of Ag-specific T cells peaked at day 5 within draining popliteal lymph nodes and 2 days later in the spleen. Preferential expression of the activation marker CD25 by tetramer-positive cells in draining popliteal nodes but not spleen suggested that gB-specific T cells were initially activated within the lymph node. In vivo cytotoxicity assays showed that Ag-specific effector cells were present within the draining lymph nodes as early as day 2 after infection, with a further 2-day lag before detection in the spleen. Consistent with the very early arming of effector CTL in the draining lymph node, adoptive transfer of CFSE-labeled gB-specific transgenic T cells showed that they had undergone one to four rounds of cell division by day 2 after infection. In contrast, proliferating T cells were first detected in appreciable numbers in the spleen on day 4, at which time they had undergone extensive cell division. These data demonstrate that HSV-1-specific T cells are rapidly activated and armed within draining lymph nodes shortly after localized HSV-1 infection. This is followed by their dissemination to other compartments such as the spleen, where they further proliferate in an Ag-independent fashion.     

Vaginal Immunization to Elicit Primary T-Cell Activation and Dissemination

Primary T-cell activation at mucosal sites is of utmost importance for the development of vaccination strategies. T-cell priming after vaginal immunization, with ovalbumin and CpG oligodeoxynucleotide adjuvant as model vaccine formulation, was studied in vivo in hormone-synchronized mice and compared to the one induced by the nasal route. Twenty-four hours after both vaginal or nasal immunization, antigen-loaded dendritic cells were detected within the respective draining lymph nodes. Vaginal immunization elicited a strong recruitment of antigen-specific CD4⁺ T cells into draining lymph nodes that was more rapid than the one observed following nasal immunization. T-cell clonal expansion was first detected in iliac lymph nodes, draining the genital tract, and proliferated T cells disseminated towards distal lymph nodes and spleen similarly to what observed following nasal immunization. T cells were indeed activated by the antigen encounter and acquired homing molecules essential to disseminate towards distal lymphoid organs as confirmed by the modulation of CD45RB, CD69, CD44 and CD62L marker expression. A multi-type Galton Watson branching process, previously used for in vitro analysis of T-cell proliferation, was applied to model in vivo CFSE proliferation data in draining lymph nodes 57 hours following immunization, in order to calculate the probabilistic decision of a cell to enter in division, rest in quiescence or migrate/die. The modelling analysis indicated that the probability of a cell to proliferate was higher following vaginal than nasal immunization. All together these data show that vaginal immunization, despite the absence of an organized mucosal associated inductive site in the genital tract, is very efficient in priming antigen-specific CD4⁺ T cells and inducing their dissemination from draining lymph nodes towards distal lymphoid organs.     

Hymenolepis nana: adoptive transfer of protective immunity and delayed type hypersensitivity response with mesenteric lymph node cells in mice

A marked degree of footpad swelling was observed in BALB/c mice infected with Hymenolepis nana eggs, when soluble egg antigen was injected into their footpads 4 to 21 days after the egg infection, indicating delayed type hypersensitivity responses in infected mice. Adoptive transfer with mesenteric lymph node cells from donor mice (BALB/c strain; +/+) infected with eggs 4 days before cell collection could confer this hypersensitivity to recipient nude mice (BALB/c strain; nu/nu). These mesenteric lymph node cells were then divided into two fractions, blast-enriched and blast-depleted cells, by density gradient centrifugation with Percoll. The recipients intravenously injected with the blast-depleted cell fraction showed a marked increase in footpad thickness, whereas the intravenous transfer of the blast-enriched cell fraction resulted in an insignificant increase in footpad thickness. The transfer of the blast-enriched cell fraction, but not of the blast-depleted cell fraction, conferred a strong adoptive immunity on syngeneic recipient nude mice, when the immunity transferred was assessed by examining cysticercoids developed in the intestinal villi on Day 4 of challenge infection. The lack of delayed type hypersensitivity response in mice that received the blast-enriched cell population was not due to a lack of the capacity of the cells to induce the response, because the cells were capable of inducing a significant increase in thickness of footpads of normal mice when these cells were locally injected into the footpad together with soluble egg antigen.(ABSTRACT TRUNCATED AT 250 WORDS)     

Recruitment of memory B cells to lymph nodes remote from the site of immunization requires an inflammatory stimulus

Successful recall Ab responses require recruitment of quiescent memory B cells to secondary lymphoid organs. However, the cellular dynamics of memory cells responding to local antigenic challenge at lymphoid sites distal from the initial Ag encounter are not well understood. We show in this study that memory B cells generated following s.c. immunization in one footpad generate secondary responses to soluble Ag given i.p. but not to Ag given s.c. in the contralateral footpad unless LPS is coadministered. Memory B cells do not express CD62L, and CD62L(-ve) cells cannot enter lymph nodes unless LPS-mediated inflammation is induced there. Functional TLR4 is required on the B cells, as well as on non-B cells, in the lymph node to achieve full recruitment. Furthermore, splenectomized mice fail to respond to such inflammatory s.c. challenge in contralateral footpads, unlike lymphadenectomized mice lacking the original draining lymph nodes. Splenectomized mice also fail to respond to i.p. challenge with soluble Ag. Together, these data indicate that, unlike the central memory pool of T cells, which circulates through resting lymph nodes, the majority of long-lived memory B cells are spleen resident and require inflammatory signals for mounting recall responses at distal challenge sites.     

Appendix and γM-antibody formation: VII. Rosette-forming cells in rabbits immunized with sheep erythrocytes

After intravenous immunization with sheep red blood cells the rabbit spleen shows a sharp rise in the number of plaque-forming cells but there is no detectable rise in PFC in the appendix or mesenteric lymph node of the same animals. Repeated immunization via an appendicostomy blind loop results in virtually no local PFC and only a small rise in splenic PFC.In lethally irradiated animals neither thymocytes nor appendix cells alone restore the splenic PFC response. Simultaneous injection of the two cell types restores both direct and indirect plaque formation. The injected cells were labeled with tritiated adenosine and a standard rosette assay for specific antigen-binding cells applied to recipients' spleen cells following immunization. Rosettes appeared by 3 days after immunization whether thymocytes or appendix cells were labeled. Labeled rosettes were observed only in animals receiving labeled appendix cells.This result demonstrates the presence of rosette forming cell precursors in rabbit appendix cell populations and suggests that the cells of gut-associated lymphoid tissues include antibody-forming cell precursors which are normally seeded to the spleen and draining lymph nodes.     

Homing of antigen-specific T cells in the Lewis rat. I. Accumulation of antigen-reactive cells in the perithymic lymph nodes

A permanent ovalbumin-specific T cell line of "helper/inducer" cell phenotype (W 3/13+, W 3/25+, OX 8-) was used to study the homing pattern in normal untreated Lewis rats. After i.v. injection, the migration of these cells was followed directly by using 51Cr- or [14C]thymidine-labeled cells. In addition, I tried to retrieve the cells from different lymphatic tissues by antigen restimulation. I found that most of the radiolabeled cells migrate to the lung, liver, kidney, and spleen. Other lymphoid tissues such as the thymus and the cervical and mesenteric lymph nodes were almost devoid of such cells with one exception: the perithymic lymph nodes (pt-LN). Twenty-four hours after the cell transfer, viable antigen-specific cells could be recovered from these organs. Within 9 days the pt-LN enlarged, the percentage of W 3/13+ and W 3/25+ T lymphocytes was enhanced, and both relatively high spontaneous and antigen-driven responses were measurable in cell cultures of these lymph nodes. All the effects were observed if viable but not irradiated antigen-specific T blasts were transferred. Moreover, after active immunization, antigen-reactive cells appeared to accumulate not only in the draining but also in the pt-LN. In both experimental situations, the adoptive transfer and the in vivo activation of antigen-specific lymphocytes, the pt-LN appear to play an important role in the homing of such cells.     

Relationship of germinal centers in lymphoid tissue to immunologic memory. VI. Transfer of B cell memory with lymph node cells fractionated according to their receptors for peanut agglutinin

We isolated germinal center B cells by exploiting their high affinity for peanut agglutinin (PNA). The PNA+ and PNA- B cells, fractionated by panning on PNA-coated petri dishes, were examined for their ability to transfer memory responses to irradiated recipients at various times after priming. With such fractionated B cells from lymph nodes taken at the peak of germinal center formation, the largest response was obtained in recipients of the PNA+ B cell population. At 4 to 5 wk after priming, and 10 days after challenge with an unrelated antigen, memory responses were approximately equal in recipients of PNA+ or PNA- B cells. At 14 wk after priming, memory responses were found only in recipients of the PNA- B cell population. Memory B cells from the spleen, taken from mice primed in the footpad 8 wk earlier, were also PNA-. Finally, we show that boosting with a TNP-conjugate in the footpad, 6 mo after priming in the same footpad, induced the reappearance of marked memory responsiveness in the PNA+ B cell fraction of the draining node.     

Antigen specific lymphocyte transformation, delayed hypersensitivity and protective immunity. I. Kinetics of the response

The association between protective immunity, delayed hypersensitivity (DTH) and in vitro antigen specific lymphocyte transformation (ASLT) has been studied during the course of BCG infection in mice. The peak ASLT response was obtained 2 weeks after a subcutaneous footpad infection using lymph node cells draining the immunization site. This response decreased to a low but statistically significant level by week 4 and persisted indefinitely thereafter. A similar course of events was observed in splenic cell cultures, but the size of the response was smaller. The reactive cells were shown to be T-lymphocytes. The ability to express DTH to tuberculin purified protein derivative coincided with the emergence of cells capable of responding in ASLT assays. In contrast, there was a poor association between ASLT and protective immunity: lymphocytes from lymph nodes that did not drain the immunization site conferred substantial protective immunity but nonetheless failed to respond significantly in ASLT assays. The latter failure could not be attributed to insufficiency of sensitized lymphocytes and was independent of the antigen concentration and the culture incubation time. It was concluded that the ASLT reactive cells were either immunoblasts or recent progeny of these cells.     

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.     

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.     

Maturation of b lymphocytes in rates. III. Two subpopulations of memory B cells in the thoracic duct lymph differ by size, turnover rate, and surface immunoglobulin.

We examined the ability of large and small thoracic duct cells obtained from Lewis rats primed to DNP to restore the adoptive secondary anti-DNP response in irradiated syngeneic hosts given an excess of T helper cells. The large cells were four to five times more active on a per cell basis than were the small cells. However, the large cells constitute only 7 to 8% of the thoracic duct cells and, therefore, make a minor contribution to the restorative activity of the unfractionated cells. Pretreatment of thoracic duct cell donors with high specific activity 3H-TdR for 48 hr before cannulation markedly reduced the memory B cell activity of the large cells, but had little effect on that of small cells. In addition, the activity of the large cells diminished with time after primary immunization, but that of the small cells remained stable. Immunofluorescent staining of the large and small cells for surface IgM and IgG, and subsequent sorting on the fluorescence-activated cell sorter (FA CS), showed that surface IgM was present on large memory B cells, and that IgG was present on small memory B cells. The experimental results suggest that two subpopulations of memory B cells in the thoracic duct lymph differ by size, rate of turnover, persistence after primary immunization, and class of surface IgG.     

Memory B cells express a phenotype consistent with migratory competence after secondary but not short-term primary immunization

The cell surface phenotype of dinitrophenol (DNP)-specific memory B cells, defined by their capacity to transfer IgG responses into syngeneic irradiated recipients, was assessed using two markers of relevance to lymphocyte migratory properties: (i) peanut agglutinin, which binds to terminal galactosyl residues expressed at high levels by several nonmigrating lymphocyte subsets and, among lymph node B cells, is highly specific for germinal center cells; and (ii) MEL-14, a monoclonal antibody specific for lymphocyte surface receptors required for migration from the blood into peripheral lymph nodes. At various times after primary or secondary immunization with DNP-keyhole limpet hemocyananin (KLH), lymph node B cells were separated by fluorescence-activated cell sorting on the basis of staining with PNA and/or MEL-14, and the presence of B-memory cells in each fraction was assessed by adoptive transfer with antigen (DNP-KLH) and helper T cells. One week after immunization, most of the memory sorted in the PNAhi population, confirming a previous report by R. F. Coico, B. S. Bhogal, and G. J. Thorbecke (J. Immunol. 131, 2254, 1983) that early memory B cells or their precursors are contained within the germinal center cell population, a population which is known to be MEL-14- and migratory-incompetent. Six weeks after primary stimulation, however, the bulk of memory cells, unlike germinal center cells, were MEL-14hi. After secondary immunization, memory was still predominantly MEL-14+ and PNAlo, although in some experiments adoptive responses were transferred by all sorted fractions. The results are consistent with the hypothesis that antigen-specific B cells initially undergo local (sessile) differentiation and proliferation in germinal centers, where they develop the capacity for adoptive transfer of antigen-specific secondary responses, but that with continued development their long-lived memory-containing progeny express a phenotype permitting their reentry into the recirculating lymphocyte pool.     

Antigen-initiated B lymphocyte differentiation. IX. Characterization of memory AFC progenitors by buoyant density and sedimentation velocity separation.

The characteristics of memory B cell antibody-forming cell (AFC) progenitors from long-term hapten-primed CBA mice were investigated by using sedimentation velocity and buoyant density separation to isolate physically distinct B cell sub-sets. The isolated fractions were assayed by the adoptive immune response to NIP-POL antigen, under conditions where neither T cells nor other accessory cells were limiting the IgM or IgG AFC responses. The results were compared to previous studies on the IgM AFC-progenitors of unprimed adult mice. Splenic IgM and IgG memory AFC-progenitor activity was largely found among the typical B cells of slow to medium sedimentation rate, in contrast to the fastre sedimenting IgM AFC-progenitor activity of unprimed animals. Splenic IgM and IgG memory AFC-progenitor activity was found among the medium to light density cells, and so resembled by this parameter the IgM AFC-progenitor activity in unprimed animals. Thoracic duct lymphocytes from hapten-primed mice also exhibited memory IgM and IgG AFC-progenitor activity in the slow-medium sedimentation range. However, in contrast to spleen, the IgM and IgG memory AFC-progenitor activity in lymph was found among very dense B cells. Two physically distinct sub-populations of memory B cells have thus been identified, namely: i) small, medium-light density, presumably tissue-resident B lymphocytes found in spleen; and ii) small, dense, presumably recirculating B lymphocytes found in lymph. Both physical forms include IgM and IgG progenitors. Both forms are distinct from the larger, medium-light density "virgin" AFC-progenitors in the spleen of unprimed adult mice.     

Murine TH response to influenza virus: recognition of hemagglutinin, neuraminidase, matrix, and nucleoproteins

BALB/c mice were primed with type A influenza virus by footpad injection or by aerosol infection with PR8 [A/PR/8/34-(H1N1)]. Isolated T cells from draining lymph nodes were then tested for their proliferation in the presence of purified viral proteins hemagglutinin, neuraminidase, matrix, and nucleoprotein. Significant responses [( 3H]thymidine incorporation) were seen against each of the four proteins after either priming scheme. When helper T (TH) cell clones were isolated by hybridoma formation from two different strains of mice, responsiveness (interleukin 2 production) towards each protein was against apparent. Of 12 virus-specific T cell hybridomas isolated, four responded to matrix, three to nucleoprotein, one to neuraminidase, three to hemagglutinin, and one cell was of undefined specificity. Each hybridoma was also tested for recognition of the HK virus [A/Hong Kong/1/68-(H3N2)], which differs in subtype from the priming strain. All matrix-specific cells, two nucleoprotein-specific cells, and the cell of undefined specificity were cross-reactive with HK virus. H1-subtype specificity was seen for all hemagglutinin and neuraminidase-specific cells and one of the three nucleoprotein-specific cells. Because many virus-immune TH cells recognize antigenically variable determinants, a significant fraction of TH cell function may be lost after virus evolution. When selecting priming schemes for long-term immunization against influenza, the isolated enhancement of TH cells recognizing conserved determinants on matrix and nucleoprotein may therefore be considered.     

IgE formation in the rat following infection with Nippostrongylus brasiliensis: I. Proliferation and differentiation of IgE-bearing cells

Sprague-Dawley rats were infected with Nippostrongylus brasiliensis larvae, and IgE formation was studied. Before infection, the serum IgE level was less than 0.4 μg/ml. The IgE level began to increase from the 10th day of infection, reached its maximum (50–100 μg/ml) at the 14th day and gradually declined. Reinfection of the rats resulted in an increase of the serum IgE level within 7 days. The IgE antibody response to N. brasiliensis antigens did not parallel the increase of IgE synthesis. In most animals, the antibody became detectable in the serum at the 21st day when the total IgE level already began to decrease. The animals showed a secondary IgE antibody response upon reinfection. Both mesenteric lymph nodes and spleen cell suspensions were examined for the presence of IgE-bearing cells (IgE-B cells) and IgE-forming cells by fluorescent antibody technique. The IgE-bearing lymphocytes became detectable in the mesenteric lymph nodes and spleen at the 8th day of infection. The proportion of the IgE-B cells in nonadherent cell population gradually increased and reached maximum at the 14th day; about 20% of immunoglobulin (Ig)-bearing cells in the mesenteric lymph nodes and 10% of Ig-bearing cells in spleen bore IgE on their surface. Evidence was obtained that these lymphocytes synthesized IgE. The IgE-forming cells were detected in both mesenteric lymph nodes and spleen of the infected animals. The number of IgE-forming cells was greater in the mesenteric lymph nodes than in spleen, indicating that the regional lymph nodes are the major source of serum IgE in the N. brasiliensis-infected animals.     

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.     

The role of dendritic cells in the initiation of immune responses to contact sensitizers. II. Studies in nude mice

Twenty-four hours after skin painting nude mice with picryl chloride, there was an increase in the number of dendritic cells (DC) isolated from the draining lymph nodes. This increased inflow or retention of DC in lymph nodes following skin painting is therefore unlikely to depend on interaction of DC with T cells. The DC obtained initiated primary proliferative responses in vitro in lymph node cells from congenic euthymic mice. Contact sensitivity developed in congenic mice when they received footpad injections of 60,000 DC from the lymph nodes of nude mice skin sensitized 1 day previously with picryl chloride or oxazolone. The initiation of delayed hypersensitivity was therefore independent of T-cell contamination within the donor DC.     

Leishmania braziliensis isolates differing at the genome level display distinctive features in BALB/c mice

Leishmania braziliensis is the species responsible for the majority of cases of human cutaneous leishmaniasis in Brazil. In the present study, L. braziliensis isolates from two different geographic areas in Brazil were studied by RAPD, using arbitrary primers. We also evaluated other biological features of these two isolates. We compared (a) the clinical features they initiate or not once delivered subcutaneously as stationary-phase promastigotes in the footpad of BALB/c mice; (b) the parasite load in both the footpad and the draining lymph node; (c) the cytokines present in the supernatant of cultures of the cell suspensions from the draining lymph nodes; and (d) the cell types present at the site of parasite delivery. The results show that the L. braziliensis strain from Ceará (H3227) is genotypically different from the L. braziliensis strain from Bahia (BA788). H3227-parasitized mice developed detectable lesions, whereas BA788-parasitized mice did not. Fifteen days post parasite inoculation there was an increase in the numbers of macrophages and lymphocytes in the footpads, whatever the parasite inoculum. Parasite load at the inoculation site--namely the footpad--did not differ significantly; in draining lymph nodes, however, it increased over the period under study. Early after parasite inoculation, the cells recovered from the draining lymph nodes of BA788-parasitized mice produced higher levels of IFN-gamma, a feature coupled to a higher number of NK cells. Later, after the parasite inoculation, there was an increased content of IL-12p70 and IL-10 in the supernatant of cells recovered from the lymph nodes of H3227-parasitized mice. This comparative analysis points out that L. braziliensis isolates differing in their genomic profiles do establish different parasitic processes in BALB/c mice.