Kenjiro Matsuno

Plasmacytoid DCs help lymph node DCs to induce anti-HSV CTLs

Antiviral cell–mediated immunity is initiated by the dendritic cell (DC) network in lymph nodes (LNs). Plasmacytoid DCs (pDCs) are known to migrate to inflamed LNs and produce interferon (IFN)-α, but their other roles in antiviral T cell immunity are unclear. We report that LN-recruited pDCs are activated to create local immune fields that generate antiviral cytotoxic T lymphocytes (CTLs) in association with LNDCs, in a model of cutaneous herpes simplex virus (HSV) infection. Although pDCs alone failed to induce CTLs, in vivo depletion of pDCs impaired CTL-mediated virus eradication. LNDCs from pDC-depleted mice showed impaired cluster formation with T cells and antigen presentation to prime CTLs. Transferring circulating pDC precursors from wild-type, but not CXCR3-deficient, mice to pDC-depleted mice restored CTL induction by impaired LNDCs. In vitro co-culture experiments revealed that pDCs provided help signals that recovered impaired LNDCs in a CD2- and CD40L-dependent manner. pDC-derived IFN-α further stimulated the recovered LNDCs to induce CTLs. Therefore, the help provided by pDCs for LNDCs in primary immune responses seems to be pivotal to optimally inducing anti-HSV CTLs.

Trafficking of host- and donor-derived dendritic cells in rat cardiac transplantation : Allosensitization in the spleen and hepatic nodes

BACKGROUND Kinetics and role of host and donor dendritic cells (DCs) in transplantation immunity are still ill-defined. Using a rat cardiac transplantation model, we studied DC trafficking and sites for allosensitization. METHODS Host and donor DCs were defined as host- or donor-type class II major histocompatibility complex antigen single-positive cells by double-immunostaining. Proliferative response of both donor and host cells were also analyzed. RESULTS Host DCs were recruited to the graft soon after transplantation. These cells represented definitive precursors because of high labeling index by a continuous bromodeoxyuridine infusion, their small round shape, and their putative bone marrow origin. Donor interstitial DCs showed a significant self-replicating capability. Both recruited host DCs in a regraft experiment and donor DCs preferentially performed blood-borne migration to the T-cell area of host spleen. Furthermore, they also migrated to the T-cell area of hepatic lymph nodes after executing the sinusoids-lymph translocation as a novel pathway for these DCs. Selectively at their migration sites, a strong T-cell proliferative response occurred, which preceded that in the graft tissues. Removal of spleen and hepatic lymph nodes significantly prolonged the mean graft survival time. CONCLUSION We conclude that allogeneic heart transplantation induces the recruitment of host DC precursors to the graft tissues and the blood-borne migration of both recruited host and donor DCs to the host spleen and hepatic nodes where effector cells are predominantly sensitized.

Splenic outer periarterial lymphoid sheath (PALS): An immunoproliferative microenvironment constituted by antigen-laden marginal metallophils and ED2-positive macrophages in the rat

SummaryIn an attempt to reveal the role of antigen-laden marginal metallophil (MM) and other macrophages in the intrasplenic immune response of a specific B-cell lineage to a thymus-independent type-2 antigen (Ficoll conjugated with fluorescein isothiocyanate), simultaneous immuno-histological observations of the involved cells were performed in the rat. By newly established methods of double or triple immunostainings, time-kinetics of the following parameters were studied and compared: (1) the antigen, (2) the specific antibody-forming cells (AFC) directed to the fluorescein-isothiocyanate determinant, (3) proliferating cells labeled with 5-bromo-2′-deoxyuridine (BrdU), and (4) macrophage subpopulations recognized by monoclonal antibodies (ED2 and ED3). The antigen localized stably not only in the marginal-zone macrophages but also in the MM except around the follicular area. The increase of BrdU-positive cells was observed from day 2 up to day 4 after antigen injection mostly in the periphery of the periarterial lymphoid sheath (outer PALS), which indicated antigen-induced proliferation. As a novel finding, the majority of AFC, both BrdU-positive and -negative, were either closely associated with the antigen-laden MM, or forming cell clusters with ED2-positive macrophages in the outer PALS. In contrast, there were very few AFC in juxtaposition to antigen-free MM in the follicular area or the antigen-laden marginal zone macrophages. The results led to the proposal of a hypothesis that the antigen-laden MM together with ED2-positive macrophages constitute an immunoproliferative microenvironment for the plasmacellular reaction by accumulating the antigen-specific B-cell lineage and promoting these cells to differentiate into the AFC and to proliferate in the outer PALS.

Effect of a single administration of testosterone on the immune response and lymphoid tissues in mice

Abstract Female mice were given a single intraperitoneal injection of testosterone immediately after irradiation and marrow reconstitution. Thirty days later testosterone had no suppressive effect on the recovery of thymus and spleen weights. Testosterone had no effect on the graft-versus-host reaction. Testosterone had no influence on the survival of the skin homografts. However, the plaque-forming cell response to sheep erythrocytes in the spleen was dramatically suppressed by testosterone. Histological observations revealed marked inhibition of lymphoid regeneration selectively in the thymus-independent areas of the peripheral lymphoid tissues. These results suggest that testosterone would act mainly on the differentiation of stem cells toward the population of bone marrow-derived B lymphocytes. The immune response to sheep erythrocytes was restored completely 90 days after testosterone administration. Testosterone given to normal adult mice can also have suppressive activity on the immune system 30 days after a single intraperitoneal injection.

Splenic marginal-zone macrophages and marginal metallophils in rats and mice

SummaryThe splenic macrophages of rats and mice were studied by light and fluorescence microscopy to determine their phagocytotic uptake of carbon and neutral polysaccharide (Fic-F), and their lysosomal enzyme activities. In rats, the large macrophages of the marginal zone (MZ) showed a moderate to strong acid phosphatase activity, and took up most of the Fic-F, even though they showed a weak phagocytotic activity to carbon particles. Red-pulp macrophages, however, ingested a large quantity of carbon particles, and are considered to be the major scavengers in the rat spleen. In contrast, the MZ macrophages in the mouse spleen were the major scavengers and showed a vigorous uptake of both carbon and Fic-F. In rats, the marginal metallophils (MM), located at the outer border of the periarterial lymphatic sheath and boundary between the MZ bridging channel and surrounding tissue, ingested Fic-F, whereas those located around the follicular area did not. In mice, on the other hand, the MM never ingested Fic-F. Lightly carbon-ladened small cells were constantly seen in the MZ of both rats and mice. They showed little acid phosphatase activity and did not ingest Fic-F. They were also present in the blood circulation.

Migration of Dendritic Cells

The migration of dendritic cells (DCs) to lymph nodes (LNs) is pivotal to the establishment of the immune response. DCs have been proved to pass through the afferent lymphatic pathway to enter LNs from the peripheral tissues after they have scanned for self or nonself antigens. In response to danger signals, both myeloid and plasmacytoid DC precursors (mDC and pDC precursors) are rapidly mobilized into the circulation. mDC precursors are recruited to inflamed tissues in response to inflammatory chemokines and then remobilized to regional LNs in response to CCL21. In contrast, pDC precursors directly transmigrate to regional LNs via high endothelial venules in a CXCL9- and E-selectin-dependent manner. Such migration is largely dependent on systemic inflammatory reactions. After accumulating in the LNs through distinct trafficking pathways, DCs interact with lymphocytes temporally and spatially to establish effective immune responses. The inflammation-dependent, chemokine-driven property of DC precursor trafficking is a very sophisticated host defense system.

Isolation of dendritic cells in the rat liver lymph.

Lymphadenectomy of the regional lymph nodes of the rat liver resulted in the direct influx of peripheral hepatic lymph into the thoracic duct after regeneration of lymphatic vessels. Thus, we could obtain the dendritic cells in the hepatic lymph by cannulating the thoracic duct. By the double immunostaining, dendritic cells in cytosmears could be easily determined as non-B, non-T, and MHC class II-positive cells. The yield of dendritic cells after enrichment by the metrizamide density gradient was about 5 x 10(5)/first 16 hr collection/rat, with viability of more than 95% and purity of more than 70%. About 80% of dendritic cells were positive for OX62, which recognized the rat dendritic cell subpopulation. They showed strong stimulating activity in the primary allogeneic mixed leukocyte reaction. The method presented here should be applicable to studies of the roles of liver dendritic cells, especially in transplantation immunity.

Kupffer Cell-mediated Recruitment of Dendritic Cells to the Liver Crucial for a Host Defense

Tissue recruitment of dendritic cells (DCs) is essential for antigen presentation. When latex particulates were injected intravenously into rats, DC precursors were recruited to the liver. Propionibacterium acnes also induced the recruitment of definite mouse DC precursors. These DCs initially showed a selective binding to Kupffer cells. In the Kupffer cell-depleted rats, DCs could neither be recruited to the liver nor adhere to sinusoidal walls. Pretreatment with varied monosaccharides in vitro showed that sugar residues consisting of N-acetylgalactosamine were necessary for this binding. Mouse DC precursors had CC-chemokine receptor 1 and 5, while granulama tissues and rat Kupffer cells expressed the corresponding chemokine, macrophage inflammatory protein-1α. Recruited DC precursors phagocytosed latex or bacteria and some of them soon translocated to hepatic nodes and induced the immune response there. We conclude that after invasion of pathogens, Kupffer cells not only scavenge them but also recruit DCs/DC precursors via chemokine- and N-acetylgalactosamine-mediated interactions. The accelerated DC traffic and the presence of blood-lymph translocation would induce rapid and efficient immune responses and thus contribute to the local defense to antigens within liver tissues as well as systemic defense to blood-borne antigens.

Differential effects of GM-CSF and G-CSF on infiltration of dendritic cells during early left ventricular remodeling after myocardial infarction.

Several lines of evidence suggest that the immune activation after myocardial infarction (MI) induces secondary myocardial injury. Although dendritic cells (DC) are potent regulators of immunity, their role in MI is still undetermined. We investigated the effect of DC modulation by CSF on left ventricular (LV) remodeling after MI. MI was induced by ligation of the left coronary artery in male Wistar rats. G-CSF (20 μg/kg/day, MI-G, n = 33), a GM-CSF inducer (romurtide, 200 μg/kg/day, MI-GM, n = 28), or saline (MI-C, n = 55) was administered for 7 days. On day 14, MI-G animals had higher LV max dP/dt and smaller LV dimensions, whereas MI-GM animals had lower LV max dP/dt and larger LV dimensions than did MI-C animals, despite similar infarct size. In MI-C, OX62+ DC infiltrated the infarcted and border areas, peaking on day 7. Bromodeoxyuridine-positive DC were observed in the border area during convalescence. Infiltration by DC was decreased in MI-G animals and increased in MI-GM animals compared with MI-C (p < 0.05). In the infarcted area, the heat shock protein 70, TLR2 and TLR4, and IFN-γ expression were reduced in MI-G, but increased in MI-GM in comparison with those in MI-C animals. IL-10 expression was higher in MI-G and lower in MI-GM than in MI-C animals. In conclusion, G-CSF improves and GM-CSF exacerbates early postinfarction LV remodeling in association with modulation of DC infiltration. Suppression of DC-mediated immunity could be a new strategy for the treatment of LV remodeling after MI.

Lymph macrophages enter the germinal center of lymph nodes of guinea pigs

To determine the fate of macrophages within the afferent lymph stream, the popliteal lymph nodes at various times (3 h to 6 months) after subcutaneous injection of india ink into the footpads of guinea pigs were examined. Two types of cells which had phagocytized india ink were observed in the germinal centers. A small number of type I phagocytes (engulfing india ink as small particles which were often found together with tingible bodies in their cytoplasm) were scattered through the germinal center. A large number of type II phagocytes (packed full of india ink) presented preferentially in the medullary portion of the germinal center, together with many pyroninophil lymphoblastoid cells. In the second experiment, the afferent lymphatics of the popliteal lymph node were ligated 15-20 min after india ink injection. Although the type I phagocytes were distributed as in the first experiment, the type II phagocytes were scanty. In the third experiment, the afferent lymphatics of the popliteal lymph node were ligated 7 days after india ink injection. The type II phagocytes disappeared rapidly from the germinal center, whereas the type I phagocytes remained and were not affected by ligature. These results suggest that the type I phagocytes are the fixed macrophages or tingible body macrophages in the germinal center, and that the type II phagocytes are the macrophages migrating from the peripheral tissues. It was also shown that many macrophages reaching the regional node via afferent lymphatics entered the germinal center through the medullary pole where the cap of small lymphocytes became thinner or disappeared.