Paul Andrews

Modulation of the immune response by tachykinins.

Neuro‐immunology is becoming an increasingly important discipline of immunology. This review has examined the immunomodulatory function of one group of neuropeptides, the TK, particularly SP and NKA. These peptides are localized in primary afferent nerves which have been shown to innervate several immune organs. In addition, binding sites for the TK have been demonstrated in thymus, spleen and lymph node. Several immune cell types also express neurokinin receptors including human circulating lymphocytes with binding to the Th/i class predominating, murine T and B cells, a human T lymphoblastoid cell line, human monocytes, rabbit polymorphonuclear leucocytes and guinea‐pig macrophages. The apposition of nerves with immune cells and receptors for neuropeptides thus produces an environment for interaction between the nervous and immune systems.

Effects of capsaicin treatment on immunoglobulin secretion in the rat: further evidence for involvement of tachykinin-containing afferent nerves

Neonatal capsaicin treatment has previously been shown to diminish the primary antibody response of adult rats to the subcutaneously administered T-dependent antigen, sheep red blood cells, as measured using a modification of the Cunningham plaque-forming cell assay technique. We have now studied the kinetics of this response in adult normal, neonatally capsaicin-pretreated and neonatally capsaicin-pretreated substance P-infused rats, and examined the effects of the tachykinin antagonist Spantide, on the plaque-forming cell response. Capsaicin pretreatment did not affect the antigen-specific plaque-forming cell response over the first 4 days following antigen injection. At days 5, 6 and 7 of the response, there was a statistically significant decrease in the number of plaque-forming cells secreting antigen-specific IgM, an effect not observed in capsaicin-pretreated rats which were given a subcutaneous infusion of substance P at the time of antigen injection. The tachykinin antagonist Spantide inhibited the plaque-forming cell response in normal rats after in vivo infusion at the time of antigen injection by more than 70%. This effect of Spantide was dose dependent, occurred with maximal effect at 10 microM, and appeared to be independent of any histamine-mediated action. The results of this study provide further evidence for a receptor-mediated immunomodulatory role of tachykinin-containing primary afferent nerves.

Tachykinin-mediated modulation of the primary antibody response in rats: evidence for mediation by an NK-2 receptor

Evidence for the involvement of primary afferent nerves and their associated neuropeptides in in vivo immunologic responses has been based on experiments in rats in which destruction of primary afferent nerves by the sensory neurotoxin capsaicin results in a diminished ability of the animal to mount a primary antibody response to sheep red blood cell (SRBC) antigen. This effect was shown to be reversed by substance P infusion immediately following antigenic stimulation. In this report we show that neurokinin A (NKA) is 12 times more potent than substance P in its capacity to reverse the effects of neonatal capsaicin pretreatment on the antibody response. Neurokinin A has a pD2 of 6.65 compared to 5.98 for substance P. In addition, NKA was more potent than substance P in reversing the effects of surgical lesions 2 days prior to antigenic stimulation. The effects of the D- and L-Pro9 analogues of [Glp6, Pro9]-SP6-11 on the plaque-forming cell response in capsaicin-treated rats provide further support for the hypothesis that the tachykinin receptor modulating the primary antibody response is an NK-2 receptor. These results demonstrate, for the first time, a role for NKA in in vivo immunomodulation.

In vivo inhibition of the rat primary antibody response to antigenic stimulation by somatostatin.

Somatostatin inhibits in vitro lymphocyte proliferative responses from a variety of species including human, mouse and rat. The immunoinhibitory effects of somatostatin are thought to involve binding to specific cell surface somatostatin receptors on immunocompetent cells. This report describes an in vivo immuno inhibitory effect of somatostatin on the rat pophteal lymph node lymphocyte primary antibody response to sheep red blood cell (SRBC) stimulation. Infusion of somatostatin immediately following SRBC injection into the hind feet of rats had a dose‐related inhibitory effect. At the highest concentration used, 10 μmol/L, the level of inhibition was similar to that previously described following neonatal capsaicin treatment of rats. This suggests that neonatal capsaicin treatment may lead to decreased primary antibody responses to SRBC by a selective effect on tachykinin containing nerves and a lesser effect on somatostatin containing nerves. The immuno inhibitory effect of somatostatin was reversed by co infusion of neurokinin A but not substance P, both of which have been shown to stimulate this response. This suggests the possibility that multiple tachykinin receptors are involved in the modulation of the SRBC primary antibody response in vivo These results present evidence for an in vivo immunomodulatory role of somatostatin.

Development of T lymphocytes within the thymus and within thymic nurse cells.

Any detailed study of T-cell development within the thymus requires, as a first step, the ability of distinguish the subpopulations of lymphoid cells which may be steps along a developmental pathway. About 95% of adult murine thymic lymphocytes can be assigned to one of four discrete subpopulations as summarized in Table 1. These major subpopulations, and many of their surface markers, have been recognized for some time, and the recent application of multiparameter flow cytometric analysis (Scollay and Shortman 1983) has merely served to emphasize how clear cut this four-way division can be. This subdivision is obtained as follows.

Inhibition of lymphocyte circulation in mice by pertussis toxin

Pertussis toxin (PT), a protein toxin of Bordetella pertussis, also called pertussigen, has a wide range of biological activities, including the induction of lymphocytosis. This phenomenon was investigated by studying lymphocyte circulation in mice. Lymph node cell suspensions were exposed to PT in vitro and then injected intravenously. A double radiolabel technique was employed, in which PT‐treated and control cells were injected into the same animals. The protocol used in these experiments was chosen to demonstrate a direct effect of PT on the injected cells. After exposure to PT in vitro, cells were profoundly excluded from lymph nodes over the succeeding six days. Entry into both mesenteric and peripheral lymph nodes, but not into the spleen was inhibited by PT, and there was an accumulation of the PT‐treated cells in the blood. Cells were excluded from the lymph nodes after treatment with as little as 2 ng/mL of PT. This dose was over two orders of magnitude lower than the threshold dose of the same PT preparation required to induce lymphocyte mitogenesis in vitro. The findings in the present communication are consistent with studies using genetically modified PT in which the ADP‐ribosylating capacity of the A‐subunit was necessary for the effect of PT on lymphocytosis.

Zonal unit-gravity elutriation. A new technique for separating large cells and multicellular complexes from cell suspensions.

A new and simple technique, zonal unit-gravity elutriation, has been devised for separating very large cells, multicellular complexes, or small organisms from suspensions consisting mainly of small cells. The separation vessel is a conical chamber with an entrance at the lower, narrower part of the cone and an exit at the upper, wider part of the cone via a dome-shaped lid. A baffle at the entrance prevents turbulence from incoming fluid. Chambers of differing widths and wall slopes are chosen depending on the sedimentation rate of the particles to be separated. A small volume of the cell suspension is placed in the chamber on the bench in a cold-room. Medium stabilized by a shallow density gradient is pumped into the base of the chamber and ascends, creating a decreasing velocity gradient. Cells sediment at unit-gravity against this ascending counterstream, and are separated into bands according to sedimentation velocity. By adjusting the flow rate of the medium, different sizes of cells can be separated. Tumor cells can be enriched, and larger blast cells can be separated from small cells in lymphoid cell suspensions. The procedure produces complete separation of thymic nurse cells (epithelial-lymphoid complexes) from free thymocytes in digested thymus suspensions and produces substantial enrichment of thymic rosettes (macrophage-lymphoid complexes). A very favorable situation for applying this technique is the isolation ofTaenia taeniaformis larvae, which can be completely purified from infected liver suspensions, representing a 4×105-fold enrichment of the parasites, with high recovery, in a single 30 min operation.

The Role of the Thymic Cortex and Medulla in T Cell Differentiation

The thymus has two main cellular zones, clearly distinguishable histologically; the major outer zone, the cortex, and the smaller central zone, the medulla. Although this morphological subdivision has been known for a long time, we still don’t know the respective roles of these two thymic compartments in the generation of peripheral T cells [reviewed in 1,2]. Our ignorance in this area is surprising since we have a lot of information concerning the nature of the lymphoid and non-lymphoid cells in the two compartments. Thus, isolated thymic lymphocytes fall into two major categories: “mature” cells binding low levels of peanut agglutinin, and expressing relatively little Thy 1 but a lot of H-2 antigen, and “immature” cells which are PNAhi, Thy 1h1 and H-2low [reviewed in 3]. The mature cells are located mainly in the medulla and hence we will call them medullary-phenotype cells, while the major population (85%) of immature cells is located in the cortex and hence these are termed cortical-phenotype cells. We use the term cortical- and medullary-phenotype because a small number of cells of either phenotype could exist in the “wrong” region and could thus be, for example, of medullary phenotype but located in the cortex. We should stress, though, that most cells do occur in the “right” region, and exceptions (for which there is little hard evidence, see below) must be few in number [see ref. 3].