Interferons are cytokines that play a complex and central role in the resistance of mammalian hosts to pathogens. Type I interferon (IFN-alpha and IFN-beta) is secreted by virus-infected cells. Immune, type II, or gamma-interferon (IFN-gamma) is secreted by thymus-derived (T) cells under certain conditions of activation and by natural killer (NK) cells. Although originally defined as an agent with direct antiviral activity, the properties of IFN-gamma include regulation of several aspects of the immune response, stimulation of bactericidal activity of phagocytes, stimulation of antigen presentation through class I and class II major histocompatibility complex (MHC) molecules, orchestration of leukocyte-endothelium interactions, effects on cell proliferation and apoptosis, as well as the stimulation and repression of a variety of genes whose functional significance remains obscure. The implementation of such a variety of effects by a single cytokine is achieved by complex patterns of cell-specific gene regulation: Several IFN-gamma-regulated genes are themselves components of transcription factors. The IFN-gamma response is itself regulated by interaction with responses to other cytokines including IFN-alpha/beta, TNF-alpha, and IL-4. Over 200 genes are now known to be regulated by IFN-gamma and they are listed in a World Wide Web document that accompanies this review. However, much of the cellular response to IFN-gamma can be described in terms of a set of integrated molecular programs underlying well-defined physiological systems, for example the induction of efficient antigen processing for MHC-mediated antigen presentation, which play clearly defined roles in pathogen resistance. A promising approach to the complexity of the IFN-gamma response is to extend the analysis of the less understood IFN-gamma-regulated genes in terms of molecular programs functional in pathogen resistance.

Cellular responses to interferon-gamma.

Primary T-cell responses in lymph nodes (LNs) require contact-dependent information exchange between T cells and dendritic cells (DCs). Because lymphocytes continually enter and leave normal LNs, the resident lymphocyte pool is composed of non-synchronized cells with different dwell times that display heterogeneous behaviour in mouse LNs in vitro. Here we employ two-photon microscopy in vivo to study antigen-presenting DCs and naive T cells whose dwell time in LNs was synchronized. During the first 8 h after entering from the blood, T cells underwent multiple short encounters with DCs, progressively decreased their motility, and upregulated activation markers. During the subsequent 12 h T cells formed long-lasting stable conjugates with DCs and began to secrete interleukin-2 and interferon-gamma. On the second day, coinciding with the onset of proliferation, T cells resumed their rapid migration and short DC contacts. Thus, T-cell priming by DCs occurs in three successive stages: transient serial encounters during the first activation phase are followed by a second phase of stable contacts culminating in cytokine production, which makes a transition into a third phase of high motility and rapid proliferation.

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T-cell priming by dendritic cells in lymph nodes occurs in three distinct phases

Lymphocytes migrate from the bloodstream by recognizing and binding to specialized endothelial cells lining the high endothelial venules (HEV) in lymph nodes and Peyer's patches. We describe here a monoclonal antibody, MEL-14, specific for a lymphocyte surface molecule that appears to mediate recognition of lymph node HEV, and to be required for lymphocyte homing into lymph nodes in vivo.

A cell-surface molecule involved in organ-specific homing of lymphocytes

Since the pioneering work of Gowans and colleagues in the 1960s,1,2 much progress has been made in understanding the pivotal role of cell migration in immunity. We now have considerable knowledge of the way in which specialized leukocytes are channeled to distinct target tissues in immune responses and inflammation (Figure 1). This review will concentrate on the migration of T cells, which are at the heart of most adaptive immune responses. Since T cells respond to pathogens only on direct contact with pathogen-derived antigen, they must migrate to sites where antigen is found. The T-cell receptor recognizes a peptide . . .

T-cell function and migration. Two sides of the same coin.

α4β7 integrin mediates lymphocyte binding to the mucosal vascular addressin MAdCAM-1

The experiments presented in this paper support the idea that the output of small lymphocytes from the thoracic duct of the rat (about 10$^{9}$/day) is normally maintained by a largescale re-circulation of cells from the blood to the lymph. It has been shown that the main channel from blood to lymph lies within the lymph nodes and that small lymphocytes enter the nodes by crossing the walls of a specialized set of blood vessels, the post-capillary venules. In order to trace the fate of small lymphocytes, cells from the thoracic duct of rats were incubated for 1 h in vitro with tritiated adenosine. This labelled the RNA of about 65% of the small lymphocytes and more than 95% of the large lymphocytes; it also labelled the DNA of a proportion of the large lymphocytes. The mixture of small and large labelled lymphocytes was transfused into the blood of two groups of rats which belonged to the same highly inbred strain as the cell donors. At various times after the transfusions the thoracic ducts in one group of rats were cannulated to determine the proportion of labelled cells which could be recovered in the lymph; at corresponding times, the rats in the other group were killed and autoradiographs prepared from their tissues to determine the location of the labelled cells. The radioactive label in the RNA of small lymphocytes was stable enough to ensure that the labelled small lymphocytes which were recovered in the lymph several days after a transfusion were those which had originally been transfused into the blood. When the thoracic duct was cannulated 20 to 27 h after a transfusion, about 70% of the labelled small lymphocytes which had been transfused into the blood could be recovered from the thoracic duct over a 5-day period of lymph collection. During the first 36 to 48 h after cannulation, while the total output of small lymphocytes was falling rapidly, the proportion of labelled cells in the lymph remained approximately constant. The pool of the animal's own cells with which the labelled cells had mixed contained between 1$\cdot $5 and 2 $\times $ 10$^{9}$ small lymphocytes; this was identified as the re-circulating pool. An autoradiographic study showed that after their transfusion into the blood the labelled small lymphocytes 'homed' rapidly and in large numbers into the lymph nodes, the white pulp of the spleen and the Peyer's patches of the intestine. The concentration of labelled cells in other tissues was trivial in comparison. Labelled small lymphocytes were seen penetrating the endothelium of the post-capillary venules in the lymph nodes within 15 min of the start of a transfusion; they were traced into the cortex of the nodes and finally into the medullary lymph sinuses. Labelled small lymphocytes did not migrate into the adult thymus but a few entered the thymus of newborn rats. It was concluded that the re-circulating pool of small lymphocytes was located in the lymphoid tissue, the thymus excepted, and that the rapid 'homing' of cells into the lymph nodes had its basis in the special affinity of small lymphocytes for the endothelium of the post-capillary venules. The interpretation of these experiments was not complicated by the presence of large, as well as of small lymphocytes in the suspensions of labelled cells which were transfused. Other experiments, in which the large lymphocytes alone were labelled with tritiated thymidine, showed that most of them migrated from the blood into the wall of the gut where they assumed the appearance of primitive plasma cells; very few divided to form small lymphocytes.

The Route of Re-Circulation of Lymphocytes in the Rat

The post-capillary venules in the lymph nodes of rats have been examined with the electron microscope. The walls of these vessels normally contain many small lymphocytes, some of which are penetrating the endothelium while others are passing through the periendothelial sheath; most of the lymphocytes lie between the endothelium and the periendothelial sheath. No other leucocyte is normally present. The evidence suggests that these venules are normally the site of a large-scale migration of lymphocytes from the blood into the lymph nodes. A study of serial sections showed that lymphocytes migrate across the vessel wall by entering the endothelial cells and traversing their cytoplasm; they do not pass through the intercellular junctions. On the other hand, in inflamed lymph nodes polymorphs and monocytes emigrate through the venules by penetrating the junctions between endothelial cells. Some of the 9large lymphocytes9 in thoracic duct lymph from normal rats were found to contain endoplasmic reticulum while others had a prominent Golgi complex.

The migration of lymphocytes through the endothelium of venules in lymph nodes: an electron microscope study

The aims of this study were (a) to find a regime of immunization with cholera toxoid in rats which would establish a high density of antitoxin containing cells (ACC) in the lamina propria of the intestine and (b) to determine the origin of the ACC. The best cellular response was achieved by a single i.p. dose of toxoid in FCA followed by an intraintestinal boost 2 wk later. ACC appeared in the thoracic duct lymph 2 days after boosting, reaching a peak of about 200,000 ACC/h at 3--4 days. This was followed by the appearance of large numbers of ACC in the intestine. The i.p. dose of toxoid by itself gave rise to very few ACC in the gut or thoracic duct lymph, but it had clearly primed the gut immune system for a secondary response. Priming was also achieved by the prolonged oral intake of toxoid. The importance of the intestinal route for boosting was shown by the failure of i.p. challenge to give an ACC response in the intestine after i.p. priming and the small response it provoked after oral priming. ACC among thoracic duct lymphocytes (TDL) and in the lamina propria contained predominantly IgA. Two observations indicated that the major source of the lamina propria ACC was from cells that emerged in the thoracic duct lymph after intraintestinal challenge. Firstly, the establishment of a thoracic duct fistula immediately before challenge prevented the appearance of ACC in the intestine. Secondly, many ACC appeared in the intestine of normal rats after the injection of TDL rich in ACC. Although homing of ACC precursors to the gut was not antigen-dependent, the distribution of ACC in the lamina propria was considerably influenced by the site of the intestinal challenge, the density of ACC being greatest at or distal to the site of injection of toxoid into the lumen of the gut.

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Cellular kinetics of the intestinal immune response to cholera toxoid in rats.

These experiments show that small lymphocytes from the thoracic duct of rats are normally a mixture of thymus-derived and marrow-derived cells, and define the traffic areas in lymphoid tissues through which the two populations recirculate.

Thoracic duct lymphocytes were labeled in vitro with uridine-3H and their histological distribution in the lymphoid tissues of normal recipients was demonstrated by radioautography. Labeled lymphocytes occupied two adjacent areas distinguished by a marked difference in the intensity of labeling; heavily labeled cells were found in thymus-dependent traffic areas of lymphocyte recirculation, while lightly labeled cells localized in the thymus-independent follicular areas around germinal centers.

A corresponding heterogeneity of uridine uptake among small lymphocytes from normal donors was demonstrated by sedimentation at 1 g ; slowly sedimenting cells incorporated little uridine and localized in follicular areas after transfusion while rapidly sedimenting cells incorporated more uridine and localized in thymus-dependent areas after transfusion.

Experimentally prepared marrow-derived small lymphocytes behaved in sedimentation studies and after transfusion like a pure population of the lightly labeled small lymphocytes in normal lymph. Artificially reconstituted mixtures of marrow-derived and thymus-derived lymphocytes were qualitatively indistinguishable from normal lymphocyte populations.

https://rupress.org/jem/article-pdf/135/2/200/1084519/200.pdf

Identification of marrow-derived and thymus-derived small lymphocytes in the lymphoid tissue and thoracic duct lymph of normal rats

The chronic drainage of lymph and cells from a thoracic duct fistula in rats results in a reduction in the weight of all the lymph nodes and of their content of small lymphocytes. The primary immune response to tetanus toxoid or sheep erythrocytes is severely depressed or abolished in such animals. This unresponsive state is related to the loss of lymphocytes from the thoracic duct fistula and, not to stress factors ensuing from the trauma of operation and restraint; it can be reversed by injecting inocula which contain almost exclusively small lymphocytes. In contrast to the severe impairment of the primary immune response in lymphocyte-depleted rats, such animals show a normal response to a second injection of tetanus toxoid. The mechanism by which small lymphocytes mediate the primary immune response is discussed.

https://rupress.org/jem/article-pdf/117/2/303/1080928/303.pdf

J. L. Gowans

Papers

The Route of Re-Circulation of Lymphocytes in the Rat

The migration of lymphocytes through the endothelium of venules in lymph nodes: an electron microscope study

Cellular kinetics of the intestinal immune response to cholera toxoid in rats.

Identification of marrow-derived and thymus-derived small lymphocytes in the lymphoid tissue and thoracic duct lymph of normal rats

The antibody response of rats depleted of lymphocytes by chronic drainage from the thoracic duct