ConA Response Research Articles

A multiplexed bead assay for profiling glycosylation patterns on serum protein biomarkers of pancreatic cancer

A multiplexed bead-based immunoassay was developed to simultaneously profile glycosylation patterns of serum proteins to investigate their usefulness as biomarkers for pancreatic cancer. The multiplex assay utilized protein-specific capture antibodies chemically coupled individually to beads labeled with specific amounts of fluorescent dye. Captured proteins were detected based on the extent and specific type of glycosylation as determined by successive binding of fluorescent lectin probes. Advantages to this technique include the fact that antibodies coupled to the beads had minimal nonspecific binding to the lectins ConA/SNA, avoiding the step of chemically blocking the antibody glycans and the bead assays were performed in a 96-well filter plate enabling high-throughput screening applications with improved reproducibility. The assay was tested with ConA and SNA lectins to examine the glycosylation patterns of α-1-β glycoprotein (A1BG) and serum amyloid p (SAP) component for use as potential biomarkers for the detection of pancreatic cancer based on the results from prior biomarker studies. The results showed that the SNA response on the captured A1BG protein could distinguish chronic pancreatitis samples from pancreatic cancer with a p-value of 0.035 and for the SAP protein with SNA, a p-value of 0.026 was found between the signal of normal controls and the pancreatic cancer samples. For the ConA response, a decline in the signal for both proteins in the serum samples was found to distinguish pancreatic cancer from normal controls and renal cell carnoma samples (A1BG, p<0.05; and SAP, p<0.0001).

The influence of leptin on the activity of lung lymphocytes under simulated microgravity

Exposure to microgravity has been implicated in the compromised immune function in space travellers, resulting in opportunistic infections, poor wound healing, and cancer. Since recent studies have suggested that leptin was capable of modulating immune responses, the purpose of this study was to examine effects of microgravity on the activation and proliferation of rat lung lymphocytes and then to examine the effects of leptin-mediated signal transduction mechanisms of lymphocyte activation in these same conditions. In control conditions (T-flasks cultured cells) leptin was not able by itself to increase lymphocytes proliferation, or induce significant increase of either IL-2 production or expression of lymphocytes activation markers, such as CD25 and CD71, while it markedly enhanced the positive effects induced on these parameters by concanavalin A (ConA). Using clinostatic rotating wall vessel (RWV) bioreactors to simulate a microgravity environment, we found that ConA responsiveness was inhibited. Moreover, under these conditions, leptin was not able to reverse these impaired functions. Accordingly with the above cited inhibitory effects exerted by the simulated microgravity environment, evidence was also obtained of defects in lymphocyte intracellular signal transduction induced by the incubation in RWV bioreactors, namely concerning decreased ConA-mediated PKC activity, and reduced expression of NF-kappaB, c-fos, and ERK1/2. Again, leptin appeared to be unable in restoring a physiologic increase of these parameters, different from what could be observed after complementation of the ConA-mediated signalling with phorbol myristate acetate, which instead demonstrated to overcome the inhibition of lymphocytes activating functions, in the presence of simulated microgravity conditions.

A novel recombinant immunotoxin with the smallest ribosome‐inactivating protein Luffin P1: T‐cell cytotoxicity and prolongation of allograft survival

In the creation of stable tolerance to MHC-incompatible allografts, reducing the large mass of donor-reactive cells via apoptosis is often required. Apoptosis induction by immunotoxins targeting surface molecules specifically presented on donor-reactive cytopathic T effector (Teff) cells is a promising strategy. Traditionally, the toxin moieties are bacterial exotoxins or plant-derived ribosome-inactivating proteins (RIPs) with large molecular size and strong immunogenicity, hence causing the problems of tissue penetration, host immune reaction and quick clearance. We have identified a novel class of small molecule RIPs (<10 kD) from the seeds of the plant Luffa cylindrica. The smallest member of this family, Luffin P1, has a molecular weight of 5226.8 Da, yet possessing a highly potent inhibitory activity on cell-free protein synthesis with IC50 of 0.88 nM. We now report a recombinant hIL-2-Luffin P1 immunotoxin, which strongly inhibited T-cell proliferation in mixed lymphocyte reaction and ConA response with IC50 of 1.8–10 nM. In vivo, hIL-2-Luffin P1 significantly prolonged the survival of major MHC-mismatched skin and kidney allografts in animal models. Thus, we demonstrate for the first time the efficacy of the smallest immunotoxin that could be further combined with other pharmacological and immunological reagents for synergistic control of pathogenic lymphocytes in immune-mediated diseases.

DNA content, chromatin supraorganization, nuclear glycoproteins and RNA amounts in hepatocytes of mice expressing insulin-dependent diabetes

Chromatin supraorganization and extensibility and nuclear glycoprotein content have been reported to change in hepatocytes from mice during development and aging, as well as under starvation and refeeding conditions. In non-obese diabetic (NOD) mice, the expression of insulin-dependent diabetes may be accompanied by metabolic changes in the liver. These changes are likely to be similar to those involved in the aging processes of non-diabetic animals. Therefore, we hypothesized that the chromatin organization, as well as the physical properties and compositions of hepatocyte nuclei would also be affected in NOD mice in the same way as those in aged non-diabetic mice. Nuclear image parameters were evaluated by image analysis of Feulgen-stained preparations. Chromatin extensibility in response to gravity was observed with polarized light after lysis and toluidine blue staining. The Con-A response of nuclear glycoproteins was evaluated with scanning microspectrophotometry. These charateristics were assessed using hepatocyte imprints from female NOD mice after a 28-day period of diabetes expression. Observations and measurements were made in comparison to healthy BALB/c mice. Total RNA amounts were determined for livers of NOD and BALB/c mice. Enhanced polyploidy levels, a decrease in chromatin higher-order packing states, an increased frequency of extended chromatin fiber formation, and deeper Con-A-responsive chromatin areas were observed in the hepatocytes of the NOD mice expressing insulin-dependent diabetes. Reduced amounts of total RNA were also found in the livers of these mice. Our findings for NOD mice expressing insulin-dependent diabetes are consistent with previously reported data for old-aged mice of the inbred strain A/Uni and may reflect changes in transcriptional activities associated with the stressful physiological demands on the liver during the expression of diabetes.

In vitro splenocyte proliferation responses of BALB/c mice to salivary gland extracts of three ixodid tick species (Acari: Ixodidae)

In vitro proliferation and cytokine production were investigated in BALB/c mice splenic cell cultures that were stimulated with concanavalin A (ConA) and lipopolysaccharide (LPS) and simultaneously exposed to salivary gland extracts (SGE) of unfed and partially fed adult ixodid ticks (Ixodes ricinus, Rhipicephalus appendiculatus, Amblyomma variegatum). Generally, tick SGE enhanced proliferation of unstimulated splenocytes and SGE of unfed ticks suppressed mitogen induced proliferation. Partially fed R. appendiculatus and A. variegatum suppressed ConA responses, while partially fed I. ricinus stimulated both ConA and LPS induced proliferation. A. variegatum and R. appendiculatus females slightly enhanced LPS responses 2 days after attachment but suppressed them at the end of the slow feeding phase. In 72 h ConA induced cell cultures, interferon gamma (IFN-γ) production was suppressed by SGE of all ticks, interleukin (IL)-10 production was enhanced by unfed I. ricinus and partially fed A. variegatum males and IL-5 production was enhanced by feeding R. appendiculatus females and A. variegatum males. The study revealed variability in the responsiveness of murine splenocytes to SGE of different ixodid tick species, whereby patterns of host immunomodulation within one tick species differed between sexes and changed during feeding.

Cellular immune responses of BALB/c mice induced by intramuscular injection of PRRSV-ORF5 DNA vaccine with different doses

BALB/c mice were immunized with 50 μg, 100 μg, 200 μg of pcDNA-PRRSV-ORF5 DNA vaccine respectively by intramuscular injection, with PBS and pcDNA3.1(+) as controls. Fluorescence activated cell Sorter (FACS) was used to detect the number of CD4+ and CD8+ T-lymphocytes. T-lymphocyte proliferation test was used to detect proliferation of the T-lymphocyte cells in peripheral blood lymphocytes of mice vaccinated with pcDNA-PRRSV-ORF5 DNA vaccine. The results showed that the difference in ConA response to T-lymphocytes in blood was highly significant between all experimental groups and the control group (P < 0.01). The number of CD4+ T-lymphocytes in experimental groups was significantly higher than that of the control group 7 d after vaccination. The number of CD8+ T-lymphocytes in the experimental groups was higher than that of the control group 28 d after vaccination. Mice immunized with a higher dose (200 μg) of DNA vaccine demonstrated higher cellular immune response than those immunized with a lower dose (100 μg, 50 μg) of DNA vaccine. The results demonstrated that pcDNA-PRRSV-ORF5 DNA vaccine could induce a good cellular immune response which may be dose-dependent.

Chromatin supraorganization and extensibility in mouse hepatocytes with development and aging

Chromatin supraorganization and extensibility, which lead to the formation of extended chromatin fibers (ECF), are affected by starvation and refeeding in adult mouse hepatocytes. It is expected that they could also change with mouse development and aging. Methods used involved topochemistry, image analysis, microspectrophotometry, gravity action, and polarization microscopy. Increased nuclear areas and Feulgen-DNA amounts with advancing hepatocyte polyploidy were found with development and aging. A slightly less packed chromatin with more heterogeneously distributed condensation levels was detected in young and old mice. Con-A responsiveness was almost absent in young mice but very deep in aged mice. ECFs formed from nuclei of adult and aged mice but not from nuclei of young mice. The frequency of ECF formation with the long lysis protocol increased with aging. In young mice, a less packed chromatin state may be associated with more intense gene activity, thus increasing the DNA-nuclear matrix interactions, and inhibiting ECF formation. Reduced DNA-nuclear matrix interactions besides defects in heterochromatin formation may induce higher ECF formation and chromatin unpackaging in old mice. We suggest that differences in Con-A staining relate to different gene activity with advancing development and aging.

T‐helper cell responses in liver transplant recipients: correlation with cytomegalovirus and other major infections

Mitogen concanavalin A (ConA) response and cytomegalovirus (CMV)-specific memory response were assessed in 24 liver transplant recipients and compared with healthy subjects. Transplant recipients as compared to healthy subjects had a lower CMV memory response at 2 weeks (P=0.023), and at 1 month (P=0.06), but a comparable response at 3 months. CMV recipient+/donor+(R+/D+) patients had the greatest increase in CMV-specific memory response at 2-3 months as compared to all other groups. Within this R+/D+ group, CMV-specific memory response was significantly more robust in patients who never had CMV infection as compared to those who developed CMV infection (P=0.035). ConA response at 2 weeks was significantly lower in patients with major infections as compared to those without them (SI 5.4 vs. 38.1, P=0.039). Thus, reconstitution of CMV-specific T-helper cell response was distinct for subsets of liver transplant recipients based on the recipient and donor CMV serostatus. Impairment in proliferative response to ConA identified a subgroup of patients with major infections after liver transplantation.

Both tissue derived and bone marrow derived stromal cells are immunoregulatory

Bone marrow derived mesenchymal stem cells have been demonstrated to have immunosuppressive effects in mice and humans. These finding suggest that they may be exploited in organ transplantation settings. Here, we investigated immunoregulatory aspects of rat intestinal derived and BM MSCs. Bone marrow was extracted from six to eight week old DA rats and plated in DMEM containing 10% Fetal Bovine Serum, 2% L-Glutamine, 1% Penicillin-Streptomycin, and 1% Sodium Pyruvate (Gibco). Every 48 hours, non-adherent cells are discarded. The immunosuppressive effects of these cells in a allogenic mixed lymphocyte reaction were investigated. Intestinal derived mesenchymal cells were obtained by enzymatic (collagenase, dispase) digestion followed by plastic adherence. Mesenteric lymph nodes from both DA and Lewis rats were extracted and passed through a metal filter to extract lymphocytes. Irradiated DA lymphocytes were used as stimulators. Lymphocytes were cultured in IMDM Media containing 1% glutamine, 1% Rat Serum, 1% Non-Essential Amino Acids, 0.1% Beta-Mercapthanol, 1% Penicillin-Streptomycin, 1% Sodium Pyruvate, 1% HEPES, and 1% vitamin (Gibco). DA MSCs were added at gradual concentrations of 105, 104, 103, 102, and 101. After a four day incubation, 1uCi of tritiated thymidine was added. Lymphocytes were also cultured with 2 mcg/ml ConA. In MLR cultures, MSC dramatically suppressed MLR responses (99%) when added at concentration of 105 MSCs. Suppression was dose dependent. In ConA cultures, MSC suppression of lymphoid cells was maximal at 105 cells. MSC suppression was not MHC restricted as Lewis derived MSCs also suppressed MLR and ConA responses (>90%). Intestinal MSCs also suppressed MLR responses (>85%). MSCs suppressed skin graft rejection in an in-vivo model. The results suggest that BM and tissue derived MSCs may have clinical utility in transplantation settings.

Modulation of human lymphocyte proliferation by salivary gland extracts of ixodid ticks (Acari: Ixodidae): effect of feeding stage and sex

Ixodid ticks remain attached to their hosts for several days to weeks. During this extended feeding process new proteins involved in the modulation of host immune responses are expressed in tick salivary glands. In our study a stimulatory or inhibitory effect of salivary gland extracts (SGE) of unfed and partially fed female Ixodes ricinus (Linnaeus, 1758), female and male Amblyomma variegatum (Fabricius, 1794) and Rhipicephalus appendiculatus Neumann, 1901 ticks on human lymphocyte proliferation induced by Concanavalin A (ConA) and phytohaemagglutinin (PHA), respectively, was investigated. SGE of all female ticks examined suppressed proliferation of ConA-induced lymphocytes; highly significant suppression was observed in the presence of unfed I. ricinus and 9-day fed A. variegatum SGE. SGE of partially fed A. variegatum and I. ricinus females suppressed PHA responses of lymphocytes. Lymphocytes showed reduced PHA and ConA responses in the presence of SGE of unfed and 2-day fed R. appendiculatus females, while SGE of 6-day fed females enhanced PHA responses, but reduced their ConA responses; generally SGE of 2-day fed females displayed the strongest inhibition. Amblyomma variegatum male SGE slightly enhanced PHA, but significantly reduced ConA responses of lymphocytes and their inhibitory effect increased during feeding. SGE of unfed and 2-day fed R. appendiculatus males enhanced PHA and ConA responses and those of 6-day fed males suppressed lymphocyte proliferation. The results suggest that (i) species- and sex-specific differences exist in the effects of tick salivary gland antigens on human lymphocyte proliferation and (ii) effect of SGE on human lymphocyte responses to mitogens varies depending on the tick feeding status.

Persistent lack of human herpesvirus-6 specific T-helper cell response in liver transplant recipients.

Specific immunologic defects predisposing to human herpesvirus-6 (HHV-6), e.g. the role of HHV-6 specific T-helper cell memory response in liver transplant recipients, have not been assessed. T-helper function (mitogen ConA response) as a marker of overall immunocompetence and T-helper response (memory response) specific to HHV-6 and cytomegalovirus (CMV) were assessed in 15 liver transplant recipients and compared with 25 healthy subjects. Samples were tested pretransplant, at 2 weeks, 1 month, 2-3 months, and 1 year posttransplantation. Stimulation index (SI) >3 was considered a positive response. Seven percent (1/15) of the transplant recipients at any time posttransplantation, as compared to 64% (16/25) of the healthy subjects, had a positive HHV-6 memory response (P = 0.00065). HHV-6-specific memory response in transplant recipients at 2 weeks (SI 1.43), 1 month (SI 1.1), and 2-3 months (SI 1.3) was significantly more suppressed than in healthy subjects (SI 17.5, P = 0.0001). Although transplant recipients as compared to healthy subjects also had a lower CMV-specific memory response posttransplant (P = 0.0439), CMV-specific memory response recovered significantly at 1 month (P = 0.03) and at 2-3 months (P = 0.027) as compared to that at 2 weeks. However, HHV-6 memory response was persistently absent up to 2-3 months with partial recovery at 1 year; 7% of the patients at 2-3 months, but 25% at 1 year had a positive HHV-6 specific memory response. Forty percent (6/15) of the patients developed HHV-6 viremia a mean of 4 weeks posttransplant. Patients with HHV-6 viremia had greater suppression of HHV-6 memory response at 1 month than those without viremia (mean SI, 0.96 vs. 1.3, P = 0.08). All but one of the patients had a positive ConA response. Prolonged suppression of HHV-6 memory response, but not overall T-helper cell function was documented and may play a role in the pathogenesis of HHV-6 infection in liver transplant recipients. Memory response to CMV after liver transplantation was significantly more robust than to HHV-6.

Genetic Analysis of Concanavalin—A Response in Broilers

Nath, M., Singh, B.P., Saxena, V.K., Singh, R.V. and Dev Roy, A.K. 2001. Genetic analysis of concanavalin-A response in broilers. J. Appl. Anim. Res. 20: 171–180. To evaluate the crossbreeding parameters in respect to in vivo mitogen response in broiler, a complete 4x4 diallel design was followed using four synthetic broiler lines namely Coloured Synthetic Male Line (CSML), White Synthetic Male Line (WSML), Coloured Synthetic Female Line (CSFL) and Naked Neck Line (NNL). The in vivo cell-mediated immune response to the mitogen concanavalin-A (Con-A) was evaluated injecting 0.1ml of Con-A (lmg Con-A in 1ml of phosphate buffer saline) inter-digitally and the foot index was calculated. The results indicated that both additive and non-additive genetic variation attributed to minor genes at many loci is important, for the genetic control of in vivo Con-A response in chicken. Total non-additive components of variance also showed significant amount of heterosis in crossbred progenies. Therefore, it may be concluded that the exploitation of non-additive component of variance is possible for improvement in cell-mediated immunity in broilers.

Proinsulin C peptide obviates sympathetically mediated suppression of splenic lymphocyte activity in rats.

To investigate whether proinsulin C peptide influences sympathetic nerve activity directly or indirectly through parasympathetic nerve activity. The proliferative response of splenic lymphocytes to Concanavalin A (ConA response) which is known to be suppressed by subjection of rats to footshock or intracerebroventricular injection of corticotropin-releasing factor through sympathetic nerve activation was measured. Effect of C peptide alone or before subjection to footshock or injection of corticotropin-releasing factor was examined. Intraperitoneal injection of C peptide was without effect on the basal ConA response, while subjection to footshock or injection of corticotropin-releasing factor lowered it. In contrast, prior injection of C peptide obviated the footshock and corticotropin-releasing factor-induced suppression of the response. When given intracerebroventricularly, C peptide was also effective at much smaller doses. Prior injection of atropine cancelled the C-peptide effects. Our results indicate that C peptide counteracts the sympathetic nerve-mediated suppression of splenic lymphocyte proliferation in an atropine-sensitive manner. Thus, C peptide probably activates the parasympathetic nervous system through the afferent mechanism, that in turn antagonizes the sympathetic nerve-mediated suppression of splenic lymphocyte functions.

Central leptin suppresses splenic lymphocyte functions through activation of the corticotropin-releasing hormone-sympathetic nervous system

Leptin is one of the key afferent signals that regulate food intake and energy expenditure by acting on specific receptors in the hypothalamus. Recently, leptin was reported to activate the peripheral immune system by acting directly on lymphocytes. To elucidate the brain-mediated participation of leptin in the modulation of peripheral immune functions, we examined the effects of intracerebroventricular (icv) injection of murine recombinant leptin on the proliferative response to Concanavalin A (ConA response) of splenic lymphocytes in rats. The ConA response of splenic lymphocytes was markedly reduced 30 min after icv injection of leptin. The suppressive effect of leptin was abolished completely either by surgical severing of the splenic nerves or by icv injection of an antibody against corticotropin-releasing hormone (CRH), but only partially by an anti-urocortin antibody. Icv injection of CRH and urocortin also suppressed the ConA response of splenic lymphocytes, and the effect of urocortin was prevented by the anti-CRH antibody, while that of CRH was not prevented by the anti-urocortin antibody. These results suggest that leptin suppresses peripheral lymphocyte functions, in contrast to the direct activating effects, indirectly through the activation of the CRH (urocortin)-sympathetic nervous system.

Taenia crassiceps cysticercosis: a role for prostaglandin E2 in susceptibility.

Several biological factors are involved in susceptibility and resistance to murine cysticercosis. A substantial body of evidence implies prostaglandins as potent regulators of immune responses during parasitic diseases. Here we evaluated the role played by prostaglandin E2 in cysticercosis. Mice were treated in vivo with prostaglandin E2 or with indomethacin (a prostaglandin E2 synthesis inhibitor) before infection. Parasite growth was enhanced by prostaglandin treatment, which provoked poor Con-A responses, low Th1-type cytokines secretion, and high levels of IL-6 and IL-10. In contrast, mice receiving indomethacin showed a reduction in parasite load parallel to a strong Con-A response and high levels of IL-2 and IFN-gamma, concomitantly with a decrease in IL-4, IL-6 and IL-10 production. Indirect in vitro studies suggest that an important source of prostaglandin E2 production could be related to host's adherent cells. However, prostaglandin E2 from parasite origin cannot be discarded.

IFN-gamma-mediated prevention of graft-versus-host disease: pharmacodynamic studies and influence on proliferative capacity of chimeric spleen cells.

Recently we demonstrated that prolonged administration of IFN-gamma prevented the development of GVHD in a MHC-mismatched murine BMT model. Treatment with IFN-gamma allowed the development of mature donor-derived allo-tolerant immunocompetent cells in complete chimeric recipients. Here we present data on the pharmacodynamics of this cytokine-mediated protection against GVHD. Treatment with 50000 U IFN-gamma twice weekly for a period of 5 weeks, starting at the day of BMT, was shown to be the optimal treatment protocol, resulting in complete prevention of GVHD-related mortality. Treatment during 1 week with a three-fold higher weekly dose of IFN-gamma (50000 U six times) did not result in significantly improved survival. The start of IFN-gamma administration was a critical factor since a delay of 3 days from the time of BMT resulted in substantial GVHD-induced mortality. Furthermore, it was shown that IFN-gamma treatment inhibited the spontaneous and Con-A-induced proliferation of T cells at 7-14 days after BMT, which is the critical period for the initiation of acute GVHD. However, long-term survivors after IFN-gamma treatment showed a recovery of immunity in contrast to long-term survivors of saline-injected animals, as tested by Con-A responsiveness. It seems that injection of high dose IFN-gamma suppresses the response of potentially alloreactive donor T cells during what normally is the initiation phase of the GVH reaction (GVHR), resulting in the abrogation of GVHD.

Thermal injury-induced immunosuppression in mice: the role of macrophage-derived reactive nitrogen intermediates.

Macrophages (Mphi) have been implicated in the suppression of lymphocyte function following thermal injury. Splenocytes isolated from C57BL/ 6NCR female mice 4-7 days after thermal injury displayed suppressed proliferative responses to Concanavalin A (ConA) and lipopolysaccharide (LPS) and high levels of reactive nitrogen intermediate (RNI) production. Inhibition of nitric oxide synthase activity with NG-monomethyl-L-arginine restored ConA responses but not LPS responses. Surprisingly, ConA-stimulated interferon-gamma (IFN-gamma) production was increased in splenocytes from injured mice. IFN-gamma contributed to the RNI-mediated immunosuppression as antibodies against IFN-gamma reduced RNI production and immunosuppression. ConA-stimulated co-cultures of splenic Mphi from injured mice and normal splenocytes produced high levels of RNI only under conditions of cellular contact and splenic Mphi from injured mice were capable of suppressing normal splenocytes responses in co-culture. These results indicate that Mphi activity and specifically RNI production contribute to the suppression of T lymphocyte function after thermal injury.

Acute cold stress induces suppressor macrophages in mice.

To elucidate mechanisms underlying acute cold stress-induced immunosuppression, functions of murine peritoneal cells of monocyte/ macrophage lineage from acute cold-stressed mice (exposed to 5 degrees C for 24 h) were investigated. Proliferative responses of spleen cells from control mice (reared at 25 degrees C) stimulated with concanavalin A (ConA) were significantly suppressed by adding peritoneal exudate cells from mice immediately after acute cold stress. The proportion of adherent cells was markedly increased in the peritoneal exudate cells from acute cold-stressed mice. These adherent cells from acute cold-stressed mice were shown to be the cells responsible for the suppressor activity for ConA responses of control spleen cells. Nonadherent cells did not suppress the ConA responses. The adherent cells in peritoneal exudate cells from control mice also suppressed the ConA responses, the inhibitory effect being considerably lower than that from acute cold-stressed mice. Addition of a nitric oxide synthase substrate analogue, NG-monomethyl-L-arginine, to the mixed cell cultures of normal spleen cells and adherent cells from acute cold-stressed mice inhibited nitric oxide release and completely abolished the suppressive effect of the adherent cells, suggesting that reactive nitrogen oxide released from the activated macrophages is apparently involved in the downregulation of proliferative responses of T cells. Thus the present findings suggest that acute cold stress induces macrophages with suppressor function and that this may contribute to the immune-suppressive state seen in spleen cells from acute cold-stressed mice.

Adrenergic suppression of peripheral blood T cell reactivity in the rat is due to activation of peripheral α2-receptors

A 20-h treatment of rats with catecholamines using s.c. implantable retard tablets markedly suppresses the in vitro reactivity of peripheral blood (PBL) T lymphocytes, provided that β-receptors are blocked with propranolol (Felsner et al., 1992). The results can be summarized as follows: (i) the suppressive effect of noradrenaline + propranolol to the concanavalin A (ConA) response of PBL was abolished by the simultaneous application of the α-blocker phentolamine. Using selective agonists, the relevant receptor was identified to belong to the α 2-subtype. (ii) The α-adrenergic suppression of the PBL T cell response was likewise observed in adrenalectomized animals, which rules out the participation of secondarily induced glucocorticoids. Furthermore, the combination of noradrenaline with the watersoluble β-blocker nadolol was equally effective to suppress the ConA response of PBL. (iii) An analogous α-mediated suppression of T cell function of PBL, but not spleen cells, was observed 1 h after i.p. treatment with tyramine, which leads to the release of endogenous noradrenaline. From these results it is concluded that the adrenergic suppression of PBL T cell functions is primarily due to the activation of peripheral α 2-receptors and that it is likewise observed under acute indirect sympathomimetic treatment.

Relationship between dehydroepiandrosterone and calcitonin gene-related peptide in the mouse thymus.

Dehydroepiandrosterone (DHEA) and calcitonin gene-related peptide (CGRP) are naturally occurring substances that are reported to have both opposing and complementary effects on immune functions. In the current study, we sought to determine how they might work together to influence the mitogen-stimulated proliferation of thymocytes. In concanavalin A (ConA)-induced thymocyte proliferation assays, CGRP and DHEA each inhibited proliferation. When the CGRP antagonist CGRP-(8-37) was added to Con A-stimulated thymocytes, the proliferative response was significantly greater than the ConA response alone across a range of ConA doses. Moreover, CGRP-(8-37) blocked the inhibitory effect of DHEA. Individually, CGRP-(8-37), CGRP, DHEA, or their combination did not stimulate thymocyte proliferation in the absence of ConA. CGRP affects the proliferation of CD4+ T cells and thus may be a regional endogenous inhibitor of the proliferation of virgin mature T cells while they remain in the thymus. Furthermore, DHEA may act via endogenous CGRP on the thymus CD4+ T cell population.