Thymocyte Function Research Articles

Signaling via the AHR leads to enhanced usage of CD44v10 by murine fetal thymic emigrants: possible role for CD44 in emigration

Signaling via the endogenous arylhydrocarbon receptor (AHR) affects proliferation, differentiation, function and gene expression of thymocytes. In the present study, we show that treatment of mouse fetal thymus lobes in organ culture (FTOC) with AHR ligands results in (a) a drastic decrease in the emigration of thymocytes in terms of numbers and types of cells, and (b) preferential emigration of CD4 −CD8 − (DN) cells expressing CD44v7- and CD44v10-containing isoforms on the cell surface. Moreover, a higher level of transcripts of various other CD44 variant isoforms (CD44v) could be detected by RT-PCR in emigrants from fetal thymi exposed to either AHR-agonist during culture. Expression of CD44v9–10-containing isoforms could be exclusively detected in DN thymic emigrants. Thus, signaling via AHR by ligands alters CD44v expression patterns in a thymocyte subpopulation. Furthermore, emigration could be decreased by the addition of anti-panCD44 antibodies to TCDD-treated FTOCs, suggesting a role for CD44 in emigration.

Neonatal Capsaicin Treatment Affects Rat Thymocyte Proliferation and Cell Death by Modulating Substance P and Neurokinin-1 Receptor Expression

Herein we provide evidence that substance P (SP) and its neurokinin-1 receptor (NK-1R) expressed on thymocytes counteract thymus depletion induced by neonatal capsaicin (CPS) treatment by affecting thymocyte proliferation and apoptotic death. SP administration reversed the CPS-mediated inhibitory effects on the total thymocyte number and subset distribution, namely CD4+ and CD4– CD8– cells, through its interaction with NK-1R as shown by concomitant NK-1R (SR140333) antagonist administration. SP-induced enhancement of thymus cellularity parallels its ability of inhibiting the thymocyte apoptotic program. Indeed, exogenously administered SP completely nullified CPS-induced apoptosis, and SR140333 abrogated the SP-mediated protective effect. SP administration also stimulated concanavalin A (Con A)-induced thymocyte proliferation of CPS-treated rats, completely reversing the CPS-induced inhibition. The SP-mediated stimulation of Con A-induced thymocyte proliferation was NK-1R dependent as shown by concomitant administration of SP and SR140333 to CPS-treated rats. Our results also demonstrate that CPS treatment induces a marked decrease of thymocyte PPT-A mRNA level and endogenous SP content as evaluated by quantitative RT-PCR, in situ hybridization and cytofluorimetric analysis. By contrast, NK-1R mRNA levels were increased in thymocytes from CPS-treated rats. Exogenous SP administration augmented PPT-A, SP and NK-1R thymocyte expression in CPS-treated rats, and this enhancement was antagonized by SR140333 administration. Overall, our results strongly suggest that the immunomodulatory effects of neonatal CPS treatment on rat thymocyte functions are dependent on vanilloid-mediated regulation of SP and NK-1R functional expression by neuronal and immune cells.

Developmental immunotoxicity of lead: impact on thymic function.

Since the potential effects of early exposure to lead on thymic functions have not been fully characterized, in this study we evaluated the capacity of lead to alter thymic function in juvenile chickens following embryonic exposure. Cornell K strain White Leghorn chicken eggs were administered lead acetate (400 microg/egg) or sodium acetate (control) on embryonic development (E12) with and without thymulin supplementation. Ex vivo production of interferon-gamma (IFN-gamma)-like cytokine by thymocytes and a delayed-type hypersensitivity (DTH) reaction were measured in the juvenile. Additionally, the effects of in vitro exposure to lead on both thymocytes and thymic stromal cells (TSCs) were evaluated. Following E12 exposure to lead, ex vivo production of IFN-gamma-like cytokine by juvenile-derived thymocytes decreased significantly compared to the control. The same effect was observed when thymocytes were directly exposed to lead in vitro and stimulated with thymic stromal supernatant. In contrast, when TSCs were exposed to lead in vitro, no change was seen in their functional capacity for promoting cytokine production. In ovo supplementation with thymulin partially reversed lead-induced DTH depression without any change in IFN-gamma-like cytokine production. Embryonic exposure to thymulin alone partially depressed the DTH response. These results suggest that lead can directly influence thymocyte function in the absence of the thymic microenvironment. Since thymulin levels may influence lead-induced immunotoxicity, embryonic endocrine status may be an important consideration. Lead exposure appears to alter thymic functions directly; however, indirect effects via endocrine factors are not precluded.

内分泌撹乱化学物質の胸腺細胞に及ぼす影響

In a previous paper we reported that endocrine disrupters (EDs) such as 4, 4'-isopropylidene diphenol (Bisphenol A: BPA), bis(2-ethylhexyl) phthalate (EHP) stimulated proliferative responses and cytokine productions of murine spleen cells in vitro. In this paper we report our studies on the effect of EDs on murine thymocytes in vitro. EDs such as BPA, EHP and diethyl stilbesterol enhanced the proliferative responses of thymocytes. The proliferative responses of in vitro precultured thymocytes with EDs were markedly enhanced by the secondary stimulation with syngeneic spleen cells as antigen-presenting cells (syngeneic mixed lymphocyte reaction; MLR), when compared with thymocytes precultured without EDs. The responder cells were T cells, and they recognized the major histocompatibility complex (MHC) class II antigens on the antigen-presenting cells. The precultured T cells with EDs were enhanced in the production of cytokines such as interleukin (IL)-3, IL-4 and interferon-gamma. These results suggest that EDs have a modulating activity on thymocyte functions.

Defective thymocyte maturation by transgenic expression of a truncated form of the T lymphocyte adapter molecule and Fyn substrate, Sin.

Adapter molecules that promote protein-protein interactions play a central role in T lymphocyte differentiation and activation. In this study, we examined the role of the T lymphocyte-expressed adapter protein and Src kinase substrate, Sin, on thymocyte function using transgenic mice expressing an activated, truncated allele of Sin (SinDeltaC). We found that SinDeltaC expression led to reduced numbers of CD4(+) and CD8(+) single-positive cells and reduced thymic cellularity due to increased thymocyte apoptosis. Because the adapter properties of Sin are mediated by tyrosine-based motifs and given that Sin is a substrate for Src tyrosine kinases, we examined the involvement of these kinases in the inhibitory effects of SinDeltaC. We found that in transgenic thymocytes, SinDeltaC was constitutively phosphorylated by the Src kinase Fyn, but not by the related kinase Lck. Using SinDeltaC and fyn(-/-) animals, we also found that the expression of Fyn was required for the inhibitory effect of SinDeltaC on thymocyte apoptosis but not for SinDeltaC-mediated inhibition of T cell maturation. The inhibitory effect of SinDeltaC on thymocyte maturation correlated with defective activation of the mitogen-activated protein kinase extracellular signal-regulated kinase. Our results suggest that the Sin mutant inhibits thymocyte differentiation through Fyn-dependent and -independent mechanisms and that endogenous Sin may be an important regulator of thymocyte development.

Kappa-opioid regulation of thymocyte IL-7 receptor and C-C chemokine receptor 2 expression.

Endogenous and exogenous kappa-opioid agonists have been widely reported to modulate the immune response. We have published results that show that the superantigen-induced proliferative response of thymocytes is inhibited by the selective kappa-opioid agonist trans-3, 4-dichloro-N-methyl-N-[2-(1-pyrolidinyl)cyclohexyl] benzeneaceamide methanesulfonate (U50,488H). Previous work has established that the kappa-opioid receptor is widely expressed within the thymus; however, little is known about the role of the kappa-opioid receptor in the function of thymocytes. In the present report, we have examined the impact of U50,488H administration on the expression of cytokines in superantigen-stimulated thymocytes by RNase protection analysis. We have measured detectable levels of the cytokines IL-2, IL-4, IL-5, IL-13, and IFN-gamma, and the chemokines lymphotactin and RANTES, in stimulated thymocyte cultures; however, addition of U50,488H did not alter the expression of these cytokines. Examination of cytokine receptor expression by these thymocytes revealed a significant inhibition in the expression of the transcript for the IL-7 receptor alpha-chain (IL-7Ralpha), and these results were confirmed by flow cytometry. Surprisingly, the expression of several other cytokine receptor chains including the common gamma-chain, IL-2Rbeta, or the IL-2Ralpha, IL-4Ralpha, and IL-15Ralpha chains, was not altered. In contrast to these results, a significant elevation in the expression of the chemokine receptor CCR2 was observed in U50,488H-treated cultures. These results suggest that the kappa-opioid receptor may function to promote cellular migration at the expense of the sensitivity to the growth-promoting/maturation activity of IL-7.

The role of orphan nuclear receptor in thymocyte differentiation and lymphoid organ development.

T lymphocytes differentiate in the thymus through several phenotypically distinct stages that are tightly regulated by multiple nuclear transcription factors. Immature CD4+CD8+ double positive (DP) thymocytes make up a majority of the population in the thymus, and exhibit several phenotypic features distinct from mature T cells. DP thymocytes express only about 10% of surface TCR that are found on mature T cells and do not proliferate and produce IL-2 in response to stimulation. Several critical events of T lymphocyte maturation such as TCRalpha gene recombination, positive and negative selection, and CD4/CD8 lineage commitment occur around the DP stage. Recent studies from our group and others on the orphan nuclear receptor RORgamma and its thymus-specific isoform RORgammat support a critical role for this nuclear receptor in the regulation of DP thymocyte function. In addition, RORgamma is required for the development of lymph nodes and Peyer's patches.

Interactive effects of cocaine and gender on thymocytes: a study of in vivo repeated cocaine exposure

This study used a mouse model including both sexes to assess the impact of repeatcocaine exposure on the differentiation and function of T cell in thymus. Cocaine hydrochloride in0.9% saline, 5 mg or 40 mg⧹kg, was administrated by i.p. injection to C57BL⧹6 mice for 10days. Thymocytes were obtained 24 h after the 10th injection. Repeat in vivo cocaine exposureinhibited the proliferation of T lymphocytes in response to Con-A and Con-A plus anti-CD28. Theproliferation induced by IL-2 in the Con-A stimulated T blasts was attenuated in cocaine treatedmice. These effects were seen at a lower cocaine dose in female mice. The total number ofthymocytes was reduced. Although the percentage of mature thymocytes (CD4 +CD8 − and CD4 −CD8 + cells) was not altered, the absolute cell numberswere attenuated. Both percentage and absolute cell number of immature thymocytes (CD4 +CD8 +) decreased and the pre-mature (CD4 −CD8 −) cellsincreased. CD28 and CD25 expression were attenuated in Con-A stimulated thymocytes of micetreated with cocaine at 40 mg⧹kg. Interleukin 2 production was not significantly altered,however, γ-IFN production was decreased by cocaine exposure at 40 mg⧹kg. Inconclusion, cocaine exerts inhibitory effects on the function of mature thymocytes, and on thedifferentiation of thymocytes. A gender difference in response to cocaine was noted in that femalemice were more sensitive to lower dose of cocaine exposure.

Cypermethrin-induced alteration of thymocyte distribution and functions in prenatally-exposed rats

The synthetic pyrethroid insecticide, cypermethrin (50 mg/kg) was given during gestation to pregnant rats by gavage in corn oil. Prenatal cypermethrin exposure induced a significant decrease in the absolute number of all thymocyte subsets during the first 30 days after birth, being the double negative CD4 −CD8 −, single positive CD4 and CD8 T cells preferentially affected. Later on day 60 and 90 double positive CD4 +CD8 + and single positive thymocytes gradually recovered, while the total number of CD4 −CD8 − cells was increased. Moreover, thymocytes from rats prenatally exposed to cypermethrin showed an impaired ability to proliferate in response to different doses of Concanavalin A (ConA) and human recombinant interleukin-2 (hrIL-2) and to produce and/or release IL-2. Overall, our results indicate that cypermethrin administered during prenatal period can affect multiple steps in thymocyte differentiation pathways resulting in an altered cell subset distribution and an impairment of thymocyte functions.

CD10 (Endopeptidase 24.11) Is a Thymic Peptide-Degrading Enzyme Possibly Involved in the Regulation of Thymocyte Functions

Human immature thymocytes express significant levels of the CD10 (endopeptidase 24.11) cell surface antigen. We report here that IOB5, an anti-CD10 mAb, as well as the phorbol ester PMA down-regulate CD10 activity at the surface of human thymocytes. The kinetics of CD10 modulation were drastically different for both effectors, indicating different regulatory mechanisms. We also demonstrated that intact human thymocytes hydrolyze thymopentin and that CD10 significantly participates in this process. Finally, we found that thymopentin and to a lesser extent phosphoramidon, a specific endopeptidase 24.11 inhibitor, induced up-regulation of CD4 and CD8 molecules at the thymocyte cell surface. In view of these results, we suggest that down-regulation of endopeptidase 24.11 at the thymocyte cell surface might reduce its activity toward thymic factors possibly involved in the regulation of thymocyte functions.

Presynaptic receptors involved in the modulation of release of noradrenaline from the sympathetic nerve terminals of the rat thymus

In a previous study we have shown that, in response to electrical stimulation, there is a substantial release of noradrenaline (NA) from the sympathetic nerve terminals of the rat thymus which is of axonal, vesicular origin. In the present study neurochemical evidence was obtained that the release of NA is subject to presynaptic modulation. This modulation operates through stimulation of α 2B-adrenoreceptors, N-nicotinic, P 1-purinergic and prostaglandin E 2 presynaptic receptors. Through these receptors the release of NA, i.e., the message from the central nervous system to the thymus, can be affected by endogenous ligands or drugs. A novel, potent and highly selective competitive antagonist of the α 2-adrenoreceptor, CH-38083, significantly enhanced the release of NA, suggesting that its release in the thymus is under tonic inhibitory control exerted by endogenously released NA. Since adrenoreceptors on thymocytes involved in the modulation of certain thymocyte functions have recently been described, it is suggested that the presynaptic modulation of the release of NA in the thymus is involved in neuro-immune communication.

The effects of granulocyte-macrophage colony-stimulating factor on tumour-infiltrating lymphocytes from renal cell carcinoma.

It has been shown that granulocyte-macrophage colony-stimulating factor (GM-CSF) can induce specific and non-specific anti-tumour cytotoxicity and also stimulates the proliferation and function of peripheral lymphocytes and thymocytes. GM-CSF and interleukin 2 (IL-2) act synergistically on peripheral lymphocytes for the induction of a highly effective cytotoxic cell population. Thus, the goal of our investigation was to study the effects of GM-CSF upon expansion, proliferation and in vitro killing activity of tumour-infiltrating lymphocytes (TILs) from renal cell carcinoma (RCC). TILs from seven consecutive tumours were cultured with GM-CSF (500 or 1000 nmol ml-1) without IL-2 supplementation, with suboptimal doses of IL-2 (8 and 40 U ml-1) plus GM-CSF (1000 nmol ml-1), and with a dose of IL-2 (400 U ml-1) which sufficed alone to induce TIL development plus GM-CSF (500 or 1000 nmol ml-1). GM-CSF alone or together with suboptimal doses of IL-2 was not able to induce or facilitate TIL development in these cultures. When GM-CSF at both concentrations studied was added to optimal doses of IL-2 the resulting TIL populations proliferated significantly better and faster (+66%), resulting in a higher cell yield (+24%) at the time of maximal expansion of the TIL cultures. The length of the culture periods of TILs was not affected by GM-CSF when compared with the control cultures supplemented with IL-2 alone. In vitro killing activity of TIL populations stimulated with IL-2 and GM-CSF remained unspecific, but lysis of the autologous tumour targets as well as the allogeneic renal tumour targets was significantly enhanced (+138%) as compared with the corresponding control TILs stimulated with IL-2 alone. Lysis of the natural killer (NK)-sensitive control cell line K562 and the NK-resistant Daudi cell line remained unchanged even though FACS analysis of TILs cultured with IL-2 and 1000 nmol of GM-CSF demonstrated a significantly higher proportion of cells expressing the CD56 molecule (+50%). Specific receptors for GM-CSF could not be demonstrated on TILs from RCC. Our data demonstrate that GM-CSF alters the biological behaviour of IL-2-activated TILs from renal cell carcinoma in terms of proliferation, in vitro killing activity and cell-surface molecule expression.(ABSTRACT TRUNCATED AT 400 WORDS)

Thymic selection and thymic major histocompatibility complex class II expression are abnormal in mice undergoing graft-versus-host reactions.

The graft-vs.-host reaction (GVHR) results in damage to the epithelial and lymphoid compartments of the thymus and thus in abnormal maturation and function of thymocytes in mice undergoing GVHR. In this report, the effects of GVHR on thymic T cell receptor (TCR) expression and usage have been investigated. GVHR was induced in unirradiated F1 hybrid mice by the intravenous transfer of parental lymphoid cells. Expression of the CD3/TCR complex on thymocyte subsets defined by CD4 and CD8 was studied by three-color flow cytometry. The level of CD3/TCR was decreased on CD4+CD8-, but not CD4-CD8+, mature thymocytes. The lack of upregulation of CD3/TCR on CD4 single-positive thymocytes, but not on their CD8+ counterparts, suggested an abnormality of class II major histocompatibility complex (MHC) expression in the thymuses of mice undergoing GVHR. Immunofluorescence staining of thymic frozen sections revealed that MHC class II expression was dramatically decreased in GVH-reactive mice. GVHR-induced changes in positive and negative selection were evaluated by determining the incidence of specific V beta TCR segment usage in the thymus. In normal mice, thymocyte usage of any given V beta segment was highly consistent between individuals of the same strain and age; however, a marked divergence in the incidence of TCR V beta 6hi and V beta 8hi cells between GVH-reactive littermate mice was observed, suggesting that thymic positive selection had become disregulated in these animals. Furthermore, negative selection was defective; the incidence of phenotypically self-reactive V beta 6hi T cells was significantly greater in the thymuses of GVH-reactive mice bearing the endogenous superantigen Mls-1a than in untreated controls. Thus, mice undergoing GVHR showed defective TCR upregulation on CD4+CD8- thymocytes and changes in TCR usage reflecting aberrant thymic selection, in conjunction with decreased expression of MHC class II. Most abnormalities of TCR expression and usage on CD4+ thymocytes observed in GVH-reactive mice were analogous to those of class II knockout mice.

Multiple effects of staurosporine, a kinase inhibitor, on thymocyte functions: Comparison with the effect of tyrosine kinase inhibitors

The effects of staurosporine, a protein kinase inhibitor, on the signal transduction and proliferation of thymocytes were studied. Signal transduction in response to Concanavalin A (Con A) as well as Concanavalin A (Con A)-induced augmentation of [ 3H]inositol incorporation into phospholipids were inhibited by staurosporine (≥10 −8M). Staurosporine inhibited thymocyte proliferation in response to Con A in the presence or absence of the phorbol ester, phorbol myristate acetate (TPA) (10 nM). This inhibition was observed regardless of whether staurosporine was added together with Con A or 3 hr later. High concentrations of staurosporine (>10 −8 M) inhibited thymocyte proliferation induced by the calcium ionophore A23187 and the phorbol ester TPA, whereas lower concentrations of the inhibitor (≤10 −8M) enhanced thymidine incorporation in response to these activators. This dual effect of staurosporine was also observed in the presence of the staurosporine-related kinase inhibitor, K252a. In contrast, the tyrosine kinase inhibitor, tyrphostin AG490, inhibited the response to A23187 and TPA at all concentrations of the inhibitor and no augmentation was seen. Interleukin 2 (IL-2)-driven mitogenesis in IL-2-dependent cells was also inhibited by staurosporine. We suggest that the inhibition of thymocyte proliferation by staurosporine results from inhibition of both protein kinase C and tyrosine kinase: the augmentation of the response to A23187 and TPA results from inhibition of protein kinase C. Inhibition of signal transduction as well as inhibition of IL-2-driven mitogenesis result from inhibition of tyrosine kinase.

Maturing thymocytes in accelerated rejection of cardiac allografts in presensitized rats.

LBNF1 cardiac allografts are rejected within 36 hr in LEW rats sensitized with BN skin grafts 7 days earlier (acute rejection in unmodified hosts = 8 days). We have studied and compared the function and migration patterns of thymocytes one day after engraftment in sensitized recipients, unmodified hosts, and normal naive rats. Thymocytes from animals experiencing accelerated rejection were more mature and functionally active, as shown by a significant elevation in percentage of OX-44+ (CD37+) cells, increased alloreactivity to BN and WF antigens, and proliferative responses to Con A and exogenous IL-2. However, the cells could neither lyse BN targets in vitro nor trigger rejection of otherwise indefinitely functioning test cardiac allografts in immunologically unresponsive T cell-deficient (B) rats after adoptive transfer. The traffic of 111In-labeled thymocytes was then evaluated. The migration index increased significantly during accelerated graft rejection, with thymocytes preferentially circulating in the blood, penetrating peripheral lymph nodes--and, interestingly, migrating back to the thymus. Thus, immunoresponsive and functionally active thymocytes, which lack the ability to recognize primed specific antigen, appear during accelerated rejection of cardiac allografts in sensitized rats. These cells migrate to the periphery, and then return in large numbers to their site of origin, the thymus. Hence, this study describes a novel behavior of thymocytes in the state of host alloreactivity that is distinct from the physiological one in otherwise normal thymus.

Androgen Deprivation Induces Phenotypic and Functional Changes in the Thymus of Adult Male Mice*

The functional immunological consequences of thymic regeneration after castration were studied in adult male C57Bl/6 mice. Phenotypic profiles of thymocytes present in the enlarged thymuses of castrate animals demonstrated a significant decrease in the proportion of thymocytes positive for the suppressor/cytotoxic phenotype (CD4-CD8+; P = 0.005). Thymic enlargement in castrate animals was accompanied by increased capacity of thymocytes to incorporate thymidine in response to Concanavalin A in vitro. Spleens from castrate mice also were enlarged, and in vitro generation of functional suppressor cells by splenocytes from castrate animals was decreased. Testosterone replacement resulted in thymic regression, with a shift toward expression of mature thymocyte phenotypes, a decrease in the double-positive phenotype (CD4+CD8+), and a relative predominance of the CD4-CD8+ suppressor/cytotoxic phenotype over the CD4+CD8- helper phenotype. Unstimulated thymidine incorporation by thymocytes from androgen-treated animals was decreased compared to controls (P = 0.050). Spleen size was not altered by androgen administration. These findings suggest that in the adult animal, changes in androgen status effect alterations in thymocyte phenotypic profiles and thymocyte function, with removal of androgens shifting the T cell balance toward the CD4 helper subset and administration of androgens changing the balance toward CD8 suppressor/cytotoxic T cell predominance.

Phenotypic analysis of skin infiltrates in comparison with peripheral blood lymphocytes, spleen cells and thymocytes in early avian scleroderma

University of California at Davis line 200 (UCD-200) chickens develop a hereditary connective tissue disease characterized by severe lymphocytic infiltration, vascular occlusion and fibrosis of skin and internal organs. To identify cellular immunological abnormalities in the acute inflammatory disease stage of this animal model for progressive systemic sclerosis (PSS) we investigated the phenotypic characteristics and function of peripheral blood lymphocytes (PBL), spleen cells and thymocytes in comparison with skin infiltrating cells. Immunofluorescence and immunohistochemical analysis using monoclonal antibodies revealed the overwhelming majority of skin infiltrating mononuclear cells in the deeper dermis and subcutaneous tissue to be T cell receptor α β (TcR2) +/CD3 +/CD4 +/class II + cells, a small portion (5–10%) of which were interleukin 2 (IL-2) receptor positive. In contrast, the inflammatory infiltrate in perivascular areas of the papillary dermis was constituted of mainly TcR γ δ (TcR1) +/class II − lymphocytes. Only few B cells ( T B cell ratio > 5) were detected. These diseased chickens showed significantly reduced percentages and numbers of circulating peripheral T cells exhibiting TcR1, TcR2, CD3, CD4 or IL-2-receptor, probably owing to an increased influx into lymphoid organs and affected tissues. In contrast to healthy chickens, the thymi of UCD-200 animals revealed fewer cells expressing TcR1, TcR2 and class II antigen, suggesting an altered intrathymic maturation of the T cell lineage. Functional in vitro studies showed a significantly decreased T cell mitogen-induced proliferation rate associated with a decreased capacity to produce IL-2 and to express IL-2 receptors. In contrast to the deficient in vitro IL-2 production the sera of UCD-200 chickens contained significant levels of IL-2 bioactivity. The alteration of T lymphocyte physiology in UCD-200 chickens adds, at least in part, to the parallels between this animal model and its human counterpart. These data confirm our hypothesis that the PSS-like disease of UCD-200 chickens includes a numeric and/or functional alteration of peripheral T cell subsets, especially of TcR1 positive cells, in contrast to the pronounced accumulation in the afflicted tissues.

Innervation of the rat thymus gland

Current views from different laboratories on the innervation of the thymus gland are reviewed with particular reference to the rat. Noradrenergic nerve profiles of the sympathetic nervous system have been demonstrated in the subcapsular cortex, at the corticomedullary junction and in the cortex itself, and extremely sparsely in the medulla. By following β-adrenergic receptor development in postnatal rats, it has been shown that there is a marked increase in density and morphological organization of the receptor in the medulla with the maturation of thymocyte function (monitored by measuring the proliferation response to concanavalin A) and a sexual dimorphism during the ontogeny of the receptor. Chemical sympathectomy of adult rats with 6-hydroxydopamine (6-OHDA) or guanethidine resulted in a loss of thymus weight, decreased cellularity, and increased apoptosis but a rise in the numbers of proliferating cells in the cortex. By contrast, proliferation of peripheral T cells was reduced after the use of 6-OHDA. Chemical sympathectomy also demonstrated that there were at least three nerve nets in the gland: noradrenergic neural profiles that were destroyed with both 6-OHDA and guanethidine, vasoactive intestinal polypeptide (VIP)-positive profiles that persisted, and AChE- and CGRP-positive profiles and cells that also persisted but had a different distribution to VIP-positive fibers. Some functional correlates of thymic innervation are discussed although the subject now needs to be further researched.

Intragonadal regulation of immune system functions

Immune responses within the mammalian gonads, and in particular the testis, are deficient in spite of adequate lymphatic drainage and the presence of lymphocytes and MHC II+ macrophages. There is considerable evidence from in vivo and in vitro studies that this 'suppression' of the immune system may be due, at least in part, to localized inhibition or regulation of normal lymphocyte and/or macrophage functions within the gonads. In the testis, both steroidal and non-steroidal products of the Leydig cells, including androgens, endorphins, and inhibin-related proteins, have been implicated in mediating this activity. In turn, a number of immune cell cytokines affect steroidogenic cell function in vitro. The studies described in this paper indicated that [3H]-thymidine incorporation by adult rat thymocytes in vitro was inhibited by conditioned medium collected from short-term incubations of Percoll-purified adult rat Leydig cells, but stimulated by testicular interstitial fluid and by conditioned medium collected from short-term incubations of adult rat seminiferous tubules. The factors responsible for these effects on thymocyte function appeared to be of large molecular weight, as they were retained by ultrafiltration membranes with exclusion limits of 10,000 or 30,000 daltons. It is hypothesized that an 'immunosuppressive' mechanism, principally mediated by non-steroidal factors secreted by the steroidogenic cells of the gonadal interstitial tissue, exists within the gonads in order to prevent activation of the immune system by germ cell antigens and growth factors associated with germ cell proliferation and differentiation. This mechanism probably acts in parallel with normal antigen-specific tolerance mechanisms operating at the gonadal level. As immune responses to germ cells are believed to be a significant causative factor in infertility, particularly in men, this represents an important area for further study.

Effects of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) treatment in vivo on thymocyte functions in mice after activation in vitro

Thymocytes from 15-day old C57BL/6 mice, pretreated with 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) 4 days before sacrifice, showed an earlier response and a higher maximal cell proliferation than thymocytes from control mice upon stimulation by concanavalin A in vitro. This is partly in contrast to the conclusion from earlier published studies. IL-2 content — as measured by growth of CTLL cells — was equally high in TCDD and in control cultures at day 1. At day 2, TCDD cultures had decreased dramatically in IL-2 content, possibly due to a high rate of consumption. At this point in time, the controls still contained a high concentration of IL-2, although less than at day 1. In contrast to the increased sensitivity to mitogen stimulation, thymocytes from TCDD-treated mice induced B-cells less avidly with respect to antibody production, and could also inhibit the T-cell help of thymocytes from untreated animals, a phenomenon which could be reversed if TCDD-treated thymocytes were irradiated before culture.