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PubMed1998

Galanin is a paracrine inhibitor of gonadotroph function in the female rat.

Recent evidence suggests that pituitary galanin synthesized in the lactotroph is a paracrine regulator of lactotroph proliferation and PRL secretion and that these effects are mediated via a pituitary-specific galanin receptor, GAL-R2(orig.). At this receptor subtype, the galanin fragment 3-29 is fully active, in contrast to both the cloned GAL-R1 and GAL-R2, at which this fragment is inactive. Since paracrine communication has been demonstrated between pituitary gonadotrophs and lactotrophs, we investigated the hypothesis that galanin is also a paracrine regulator of gonadotroph function. Galanin attenuated LHRH-stimulated LH release in a dose-dependent manner in monodispersed rat anterior pituitaries harvested at proestrus (LHRH 100 nM, 10.7 +/- 0.2 ng/ml(-1) x 4 h vs. LHRH 100 nM + 1 microM porcine galanin (pGal), 7.0 +/- 0.2 ng/ml(-1) x 4 h; P < 0.01; i.e. 37% reduction). Galanin had similar suppressive effects on FSH release. Galanin, also dose-dependently, attenuated the LHRH-stimulated LH release from perifused proestrous rat pituitary fragments. pGal (1 microM) reduced the stimulated LH release by 80%, [area under the curve (AUC), LHRH 100 nM, 713 +/- 149 vs. LHRH 100 nM + 1 microM pGal, 131 +/- 7 ng/min x ml(-1) x 4 h; P < 0.02]. In addition, galanin 3-29, the specific GAL-R2(orig.) receptor agonist, inhibited LHRH-stimulated LH release from perifused proestrous rat pituitary fragments [AUC, LHRH 100 nM, 642 +/- 77 ng/min x ml(-1) vs. LHRH 100 nM + pGal 1-29, 206 +/- 44 ng/min x ml(-1) (P < 0.02); and LHRH 100 nM + pGal 3-29, 310 +/- 19 ng/min x ml(-1) (P < 0.02)]. Immunoblockade with specific galanin antiserum potentiated the LHRH-stimulated release of LH by 48% from perifused proestrous rat pituitary fragments (AUC, LHRH 100 nM + galanin antiserum, 721 +/- 65 ng/min x ml(-1) vs. LHRH 100 nM alone or with nonimmune antiserum, 489 +/- 33 ng/min x ml(-1) or 545 +/- 46 ng/min x ml(-1), P < 0.05). This data suggests that galanin may act as a paracrine agent via the pituitary-specific GAL-R2(orig.) to inhibit gonadotroph function.

Animals↗

PubMed1997

Upregulation of the angiogenic factors PlGF, VEGF and their receptors (Flt-1, Flk-1/KDR) by TSH in cultured thyrocytes and in the thyroid gland of thiouracil-fed rats suggest a TSH-dependent paracrine mechanism for goiter hypervascularization.

Placenta growth factor (PlGF) and vascular endothelial growth factor (VEGF) represent two closely related angiogenic growth factors active as homodimers or heterodimers. Since goiters of the thyroid gland are extremely hypervascular, we investigated the expression of PlGF, VEGF and their receptors, Flt-1 and Flk-1/KDR, in a small panel of human goiters from patients with Graves's disease, in an animal model of thyroid goitrogenesis and in in vitro cultured thyroid cells. Here we report that the mRNA expression of PlGF, VEGF and their receptors is markedly enhanced in biopsies of goiters resected from Graves's patients. In vivo studies demonstrated that in the thyroid gland of thiouracil-fed rats, increased mRNA and protein expression of PIGF, VEGF, Flt-1 and Flk-1/KDR occurred subsequent to the rise in the serum thyroid stimulating hormone (TSH) levels and in parallel with thyroid capillary proliferation. In vitro studies confirmed the existence of such TSH-dependent paracrine communication between thyroid epithelial cells and endothelium since the conditioned medium collected from TSH-stimulated thyrocytes acquired mitogenic activity for human umbilical vein endothelial (HUVE) cells. Altogether, these data suggest that PlGF and VEGF, released by thyrocytes in response to the chronic activation of the TSH receptor pathway, may act through a paracrine mechanism on thyroid endothelium.

Animals↗

PubMed2001

Influence of neonatal estrogens on rat prostate development.

Brief exposure of rodents to estrogens during early development alters prostate branching morphogenesis and cellular differentiation in a dose-dependant manner. If estrogenic exposures are high, these disturbances lead to permanent imprints of the prostate, which include reduced growth, differentiation defects of the epithelial cells, altered secretory function and reduced responsiveness to androgens in adulthood. This process, referred to as neonatal imprinting or developmental estrogenization, is associated with an increased incidence of prostatic lesions with aging, which include hyperplasia, inflammation and dysplasia. To better understand how early estrogenic exposures can permanently alter prostate growth and function and predispose the gland to neoplasia, the effects of estrogens on prostatic steroid receptors, cell-cell communication molecules and key developmental genes were examined. Transient and permanent alterations in the expression of prostatic androgen receptors, estrogen receptors alpha (ERalpha) and beta, and retinoic acid receptors are observed. It is proposed that the estrogen-induced alterations in these critical transcription factors play a fundamental role in initiating prostatic growth and differentiation defects. Down-stream effects of the altered steroid receptor expression include disruption of TGFbeta paracrine communication, altered expression of gap junction connexin molecules and loss of epithelial cadherin on epithelial cells. Additionally, specific disruptions in the expression of prostatic developmental genes are observed in response to neonatal estrogen. An extended developmental period of hoxa-13 expression, a lack of hoxd-13 increase with maturation, and an immediate and sustained suppression of hoxb-13 was noted within prostatic tissue. A transient decrease in Nkx3.1 expression in the developing prostate was also observed. Thus subtle and overt alterations in Hox-13 and Nkx3.1 genes may be involved in the altered prostate phenotype in response to neonatal estrogen exposure. In summary, estrogen imprinting of the prostate gland is mediated through up-regulated levels of stromal ERalpha, which initiates alterations in steroid receptor expression within the developing gland. Rather than being an androgen-dominated process, as occurs normally, prostatic development is regulated by alternate steroids, including estrogens and retinoids, in the estrogenized animal. This, in turn, leads to disruptions in the coordinated expression of critical developmental genes including TGFbeta, Hox-13 genes and Nkx3.1. Since a precise temporal expression pattern of these and other molecules is normally required for appropriate differentiation of the prostatic epithelium and stroma, the estrogen-initiated disruption in this pattern would lead to permanent differentiation defects of the prostate gland. It is hypothesized that these molecular and cellular changes initiated early in life predispose the prostate to the neoplastic state upon aging.

Animals↗

PubMed1993

Possible paracrine facilitation of ventricular growth in oculo by atrial tissue.

Load-independent growth controls were investigated in embryonic atria and ventricles cultured in the anterior eye chamber of adult rats. At embryonic day 12, the mass of atria was half that of ventricles (0.11 vs. 0.24 mg), whereas after 8 wk in oculo, atrial grafts were threefold larger than ventricular grafts (2.26 +/- 0.36 vs. 0.69 +/- 0.16 mg). To examine possible paracrine communication between atria and ventricles, two grafts were cocultured in the same eye chamber. Atrial grafts cocultured with either another atrial graft or a ventricular graft did not differ in projected area or mass from atrial grafts cultured alone. Ventricular grafts cocultured with another ventricular graft did not differ in projected area or mass from ventricular grafts cultured alone. However, ventricular grafts cocultured with an atrial graft grew larger than single ventricular grafts. At 1 wk in oculo, DNA synthesis (bromodeoxyuridine incorporation) was higher in ventricular grafts cocultured with atrial grafts than in single ventricular grafts (6.84 +/- 0.88 vs. 4.40 +/- 0.10%). Thus atria may stimulate growth, including DNA synthesis, in developing ventricles via a paracrine mechanism.

Animals↗

PubMed2005

Extrasynaptic localization of glycine receptors in the rat supraoptic nucleus: further evidence for their involvement in glia-to-neuron communication.

Neurons of the rat supraoptic nucleus (SON) express glycine receptors (GlyRs), which are implicated in the osmoregulation of neuronal activity. The endogenous agonist of the receptors has been postulated to be taurine, shown to be released from astrocytes. We here provide additional pieces of evidence supporting the absence of functional glycinergic synapses in the SON. First, we show that blockade of GlyRs with strychnine has no effect on either the amplitude or frequency of miniature inhibitory postsynaptic currents recorded in SON neurons, whereas they were all suppressed by the GABA(A) antagonist gabazine. Then, double immunostaining of sections with presynaptic markers and either GlyR or GABA(A) receptor (GABA(A)R) antibodies indicates that, in contrast with GABA(A)Rs, most GlyR membrane clusters are not localized facing presynaptic terminals, indicative of their extrasynaptic localization. Moreover, we found a striking anatomical association between SON GlyR clusters and glial fibrillary acidic protein (GFAP)-positive astroglial processes, which contain high levels of taurine. This type of correlation is specific to GlyRs, since GABA(A)R clusters show no association with GFAP-positive structures. These results substantiate and strengthen the concept of extrasynaptic GlyRs mediating a paracrine communication between astrocytes and neurons in the SON.

Animals↗

PubMed2003

Intercellular interactions in the mammalian olfactory nerve.

The small, unmyelinated axons of olfactory sensory neurons project to the olfactory bulb in densely packed fascicles, an arrangement conducive to axo-axonal interactions. We recently demonstrated ephaptic interactions between these axons in the olfactory nerve layer, the layer of the olfactory bulb in which the axon fascicles interweave and rearrange extensively. In the present study, we hypothesized that the axons, which express connexins, may have another mode of communication: gap junctions. Previous transmission electron microscopy (TEM) studies have failed to demonstrate such junctions. However, the definitive method for detecting gap junctions, freeze fracture, has not been used to examine the interaxonal connections of the olfactory nerve layer. Here, we apply a combined approach of TEM and freeze fracture to determine if gap junctions are present between the olfactory axons. Gap junctions involving olfactory axons were not found. However, by freeze fracture, P faces of both the axons and ensheathing cells (glia that surround the axon fascicles) contained distinctive linear arrays of particles, aligned along the small columns of extracellular space. In axons, few intramembranous particles were present outside of these arrays. Multi-helix proteins, including ion channels and connexin hemichannels, have been shown to be visible as particles by freeze fracture. This suggests that the proteins important for signal transmission are confined to the linear arrays. Such an arrangement would facilitate ephaptic transmission, calcium waves, current oscillations, and paracrine communication and may be important for olfactory neural code processing.

Animals↗

PubMed1998

Keratinocyte growth factor expression by the bovine corpus luteum.

Communication between cells of the corpus luteum (CL) is thought to be necessary for normal luteal function. Keratinocyte growth factor (KGF) is produced by mesenchymally derived cells in numerous tissues and acts on epithelial cells. In bovine follicles, theca cells produce KGF, which can stimulate granulosa cell proliferation. Whether KGF is produced by ovarian cells after luteinization is unknown. Our objective was to determine whether KGF mRNA and protein were present in bovine luteal tissue, and if so, to determine what type(s) of luteal cells contains KGF. CL (n = 3-4/day) were obtained from specific days throughout diestrus. Presence of KGF mRNA in CL was determined using a porcine KGF anti-sense cRNA probe. Northern analyses of luteal tissue poly(A)+ RNA revealed a single transcript (approximately 2.0 kilobases), the quantity of which did not change throughout diestrus. Western analysis revealed an immunoreactive band (28 kDa) in luteal tissues and theca cell homogenates that was absent from granulosa cell homogenates. Immunocytochemistry showed KGF predominantly in theca and small luteal cells. Results indicate that bovine CL produce and contain KGF, which is primarily localized in small luteal cells. Therefore, KGF may participate in paracrine communication within the bovine CL.

Animals↗

PubMed2002

Microglia as a source and target of cytokines.

Cytokines constitute a significant portion of the immuno- and neuromodulatory messengers that can be released by activated microglia. By virtue of potent effects on resident and invading cells, microglial cyto- and chemokines regulate innate defense mechanisms, help the initiation and influence the type of immune responses, participate in the recruitment of leukocytes to the CNS, and support attempts of tissue repair and recovery. Microglia can also receive cyto- and chemokine signals as part of auto- and paracrine communications with astrocytes, neurons, the endothelium, and leukocyte infiltrates. Strong responses and modulatory influences can be demonstrated, adding to the emerging view that microglial behavior is highly dependent on the (cytokine) environment and that reactions to a challenge may vary with the stimulation context. In principle, microglial activation aims at CNS protection. However, failed microglial engagement due to excessive or sustained activation could significantly contribute to acute and chronic neuropathologies. Dysregulation of microglial cytokine production could thereby promote harmful actions of the defense mechanisms, result in direct neurotoxicity, as well as disturb neural cell functions as they are sensitive to cytokine signaling.

Animals↗

PubMed2002

The role of cytokines in the epithelial cancer microenvironment.

The epithelial tumour microenvironment is a complex tissue comprising variable numbers of tumour cells, fibroblasts, endothelial cells and infiltrating leucocytes. Cytokines are key molecules controlling autocrine or paracrine communications within and between these individual cell types. Under some circumstances, endogenous cytokines may orchestrate host responses against the tumour, but there is increasing evidence that the cytokine network contributes to tumour growth, progression and host immuno-suppression. In this review we outline some of the actions of endogenous cytokines in epithelial tumours with particular emphasis on tumour necrosis factor alpha, TNF, related inflammatory cytokines and the chemokine group of chemoattractant cytokines.

Animals↗

PubMed1991

Selectivity of juxtaposition between cup-shaped lactotrophs and gonadotrophs from rat anterior pituitary in culture.

Semi-thin sections of three-dimensional reaggregates from adult female rat pituitary, cultured in serum-free defined medium, were stained for prolactin, gonadotropin, thyrotropin, growth hormone and S-100, using the double immunolabelling technique. The frequency of juxtaposition between lactotrophs and gonadotrophs was enumerated and compared with the expected frequency at random distribution of polygonal cell profiles in a hexagonal configuration. The proportions of lactotrophs and gonadotrophs in the aggregate sections were determined using stereometrical analysis. The observed frequency of juxtaposition did not differ significantly from the expected frequency. Hence, no reason was found to assume a selective adhesion between lactotrophs and gonadotrophs in adult female rat pituitary reaggregates. A constant proportion of lactotrophs was found to meet the criteria of a cup-shaped morphology, and 70% +/- 9% (mean +/- S.D.) of these so-called cup-shaped lactotrophs were found to be juxtaposed at their concave side to gonadotrophs. Administration of 0.01 nM 17 beta-oestradiol to the culture medium resulted in a significant reduction of the proportion of cup-shaped lactotrophs but did not affect the selectivity of juxtaposition to gonadotrophs. The selectivity of juxtaposition between cup-shaped lactotrophs and gonadotrophs may be the morphological correlate of the functional relationship between these cells, which are known to be involved in an intra-pituitary paracrine communication system.

Animals↗

PubMed2000

Changes in the expression of the nerve growth factor receptors TrkA and p75LNGR in the rat thymus with ageing and increased nerve growth factor plasma levels.

The nerve growth factor (NGF) receptors p75LNGR and TrkA are expressed by thymic epithelial cells. Presumably, the NGF-TrkA system is involved in the paracrine communication between thymic epithelial cells and thymocytes, whereas the functional role of p75LNGR is still unknown. The thymus of vertebrates undergoes age-related changes that in part depend on hormonal factors. In order to find out whether thymic epithelial cells are responsive to NGF during the whole lifespan of the rat, we studied NGF receptor expression in the thymus from birth to 2 years of age, using immunohistochemistry. Furthermore, to evaluate whether increased plasma levels of NGF affected the ageing process, either NGF or 4-methylcatechol (4MC), an inductor of NGF synthesis, was administered. Both TrkA and p75LNGR were expressed by a subpopulation of thymic epithelial cells during the whole age range studied and their expression peaked at around 3 months. TrkA was primarily found in subcortical and medullary epithelial cells, whereas p75LNGR was seen in a subpopulation of medullary cells. Cortical epithelial cells, neural crest-derived cells, other stromal cells and thymocytes were not immunoreactive for NGF receptors. Neither the administration of NGF nor the increased NGF plasma levels obtained after 4MC treatment seemed to affect the ageing of the thymus as assessed by morphological and immunohistochemical criteria, but this increase in NGF levels did produce a shift in the expression of p75LNGR from epithelial cells to ED1-positive macrophages in animals of 6 months and older. Present results indicate that the expression of p75LNGR and TrkA in the rat thymus undergoes age-dependent changes that parallel those of epithelial cells. NGF could therefore be important for thymus homeostasis, possibly acting on epithelial cells. Nevertheless, NGF did not seem to be able to prevent the involution of this organ, although it produced a switch in the expression of p75LNGR, the significance of which remains to be established.

Aging↗

PubMed1995

NO in the lung.

In the lung, nitric oxide (NO) derives from several cellular sources, forming networks of paracrine communication. In pulmonary vessels, NO produced by endothelial cells is a powerful vasodilator. In the airways, NO originates from epithelial cells and from adventitial nerve endings to induce smooth muscle relaxation. Activated macrophages can also produce large quantities of NO during lung immunological reactions. In the normal pulmonary circulation, NO not only mediates vasodilation, but also opposes vasoconstriction, prevents platelet adhesion, controls growth of smooth muscle and influences the composition of the extracellular matrix. During exposure to chronic hypoxia, impaired endothelial NO production contributes to the increased vasomotor tone and vascular remodelling leading to sustained pulmonary hypertension. Exogenous NO gas delivered via the airspaces is a selective pulmonary vasodilator. Inhaled NO is now used as a therapy to treat various forms of pulmonary hypertension and to improve arterial oxygenation during lung injury.

Animals↗

PubMed1991

Secretory granule and synaptic vesicle formation.

Sorting of secreted proteins into dense-core secretory granules may involve selective aggregation of regulated secretory proteins, rather than a conventional sortase. Synaptic vesicles, which mediate paracrine communication between adjacent cells, appear to arise by a modification of the early endosome pathway. Targeting to the cell surface involves the actin-based cytoskeleton and small GTP-binding proteins.

Animals↗

PubMed1999

Vasoactive intestinal peptide neurons as synaptic targets for vasopressin neurons in the suprachiasmatic nucleus. Double-label immunocytochemical demonstration in the rat.

Cellular relationships between neurons producing vasopressin or vasoactive intestinal peptide in the suprachiasmatic nucleus of the hypothalamus, the main component of the central circadian timing system in mammals, were investigated in the rat using double immunocytochemistry. Analysis of serial confocal images revealed that the vasopressin-synthesizing neurons not only are important targets for the vasoactive intestinal peptide-synthesizing neurons, as previously demonstrated, but also establish reciprocal axosomatic contacts with these neurons, which have never been reported. On average, 5.4 vasoactive intestinal peptide contacts per vasopressin perikaryon and 1.7 vasopressin contacts per vasoactive intestinal peptide perikaryon were counted. That both types of neurons are linked by reciprocal synapses was confirmed at the electron microscopic level using a combination of immunoperoxidase and immunogold-silver labeling. Existence of an anatomical substrate for a vasopressinergic control of the vasoactive intestinal peptide neurons may have important functional consequences. In view (i) of the presumed, direct or indirect, involvement of the vasopressin neurons in relaying pacemaker information within and outside the suprachiasmatic nucleus, and (ii) of the established role of the vasoactive intestinal peptide neurons as the main light-sensitive cells, it provides support for a neuronal mechanism through which the circadian clock may regulate inputs related to environmental messages. Our electron-microscopic data also extended earlier observations, pointing to the involvement of vasopressin and vasoactive intestinal peptide terminals in so-called double synapses that, conceivably, could regulate neuronal synchronization in the suprachiasmatic nucleus. A morphological basis for non-synaptic interactions that could be involved in ephaptic and/or paracrine communication between both types of peptidergic neurons is also reported.

Animals↗

PubMed2000

Effects of cholecystokinin on acid formation in glands and cells isolated from rabbit and rat gastric mucosa.

Isolated gastric glands and isolated cells prepared from rabbit and rat were studied to analyse the influence of cholecystokinin octapeptide (CCK 8) on histamine stimulated parietal cell acid formation as assessed by [14C]aminopyrine sequestered in acid tissue compartments. In rabbit gastric glands, CCK 8 evoked 32+/-6% (P<0. 01) inhibition of histamine stimulated acid formation, whereas in glands prepared from rat no inhibition was recorded. Instead, CCK 8 seemed to induce a variable increase of the histamine stimulation in rat gastric glands as the aminopyrine accumulation was increased by 110+/-46% (P<0.1). Further studies on cell preparations derived from rabbit gastric mucosa revealed dual properties of CCK 8, eliciting either inhibition or stimulation of the parietal cell depending on the presence of endocrine cells. The results show that paracrine communication may be effective in glandular preparations, but seems to vary depending on species.

Aminopyrine↗

PubMed1999

Functional characterization of N-methyl-D-aspartic acid-gated channels in bone cells.

Our recent identification of glutamate receptors in bone cells suggested a novel means of paracrine communication in the skeleton. To determine whether these receptors are functional, we investigated the effects of the excitatory amino acid, glutamate, and the pharmacological ligand, N-methyl-D-aspartic acid (NMDA), on glutamate-like receptors in the human osteoblastic cell lines MG63 and SaOS-2. Glutamate binds to osteoblasts, with a Kd of approximately 10(-4) mol/L and the NMDA receptor antagonist, D(L)-2-amino-5-phosphonovaleric acid (D-APV), inhibits binding. Using the patch-clamp technique, we measured whole-cell currents before and after addition of L-glutamate or NMDA and investigated the effects of the NMDA channel blockers, dizolcipine maleate (MK801), and Mg2+, and the competitive NMDA receptor antagonist, 3-((R)-2-carboxypiperazin-4-yl)-propyl-1-phosphoric acid (R-CPP), on agonist-induced currents. Both glutamate and NMDA induced significant increases in membrane currents. Application of Mg2+ (200 micromol/L) and MK801 (100 micromol/L) caused a significant decrease in inward currents elicited in response to agonist stimulation. The competitive NMDA receptor antagonist, R-CPP (100 micromol/L), also partially blocked the NMDA-induced currents in MG63 cells. This effect was reversed by addition of further NMDA (100 micromol/L). In Fura-2-loaded osteoblasts, glutamate induced elevation of intracellular free calcium, which was blocked by MK801. These results support the hypothesis that glutamate plays a role in bone cell signaling and suggest a possible role for glutamate agonists/antagonists in the treatment of bone diseases.

Binding Sites↗

PubMed2000

Immunohistochemical study of the glutamate transporter proteins GLT-1 and GLAST in rat and gerbil pineal gland.

Several investigations performed during this decade have led to the hypothesis that small secretory vesicles of pinealocytes (generally referred to as synaptic-like microvesicles, SLMVs) are components of a system for intercellular paracrine communication between pineal cells, which shares many features with the process of synaptic neurotransmission. According to a recent study, one parallel that can be drawn to synaptic signal transduction seems to be the presence of pineal re-uptake systems for messenger molecules released from SLMVs, i.e. for neuroactive amino acids such as L-glutamate. In order to further characterize these uptake mechanisms, we have carried out an immunohistochemical study to explore the presence and cellular localization of the glutamate transporters GLT-1 and GLAST in rat and gerbil pineal glands. GLT-1 and GLAST were always detected in a subpopulation of pineal parenchymal cells in both species. Using immunostaining of serial semithin sections with antibodies against marker proteins of pineal cell types, GLT-1- and GLAST-positive cells were identified as interstitial glial cells. In addition, some pinealocytes also displayed immunoreactivity for GLT-1. In contrast to current thinking, our findings show that GLT-1 is not the only glutamate transporter subtype expressed in the pineal gland. Moreover, our observations point to a significant participation of interstitial cells in the process of pineal glutamatergic communication, reminiscent of the role of glial cells during glutamatergic neurotransmission in the central nervous system.

ATP-Binding Cassette Transporters↗

PubMed1997

Heterogeneity of pituitary folliculo-stellate cells: implications for interleukin-6 production and accessory function in vitro.

The population of folliculo-stellate (FS) cells of the rat anterior pituitary has been shown to be ultrastructurally and immunohistochemically heterogeneous. Based on the overlap of ultrastructural characteristics, the localization in the anterior pituitary and the co-expression within the same cel of the S-100 protein (a marker for FS cells) and MHC-class II determinants (an immune marker) we concluded that a partial overlap exists between the population of FS cells and the monocyte-derived dendritic cells (DC). In this report we describe that interleukin-6 (IL-6) immunoreactivity was found in situ in stellate cells of the rat, mouse and human anterior pituitary at a very low density (< 1% of the cells); the topography was reminiscent of the distribution of FS cells. In the present study we also analyse three different pituitary cell separation methods, in order to study the functional heterogeneity of the FS cells in vitro, and to verify whether functionally distinct subpopulations exist within the FS cell group. Production of bioactive IL-6 was measured in conditioned media of rat anterior pituitary cells separated by (i) bovine serum albumin (BSA) gradient sedimentation at 1 g, (ii) Nycodenz gradient and (iii) a magnetic cell separation (MACS) technique. Production of bioactive IL-6 by cell cultures of 1 to 4 days was correlated with the proportional number of S100 immunoreactive and S100 producing cells, but was not correlated with the proportional number of MHC-class II expressing (OX6-positive) dendritic cells (DC). The distribution pattern of OX6-positive DC was found to partly overlap with the distribution pattern of S100-positive cells in the BSA gradient. Co-sedimentation of S100-positive FS cells and MHC-class II-expressing DC was not restricted to the top fractions of the BSA gradient, but was also found in the low density Nycodenz fraction. MACS separation of the rat anterior pituitary cells resulted into a population enriched in OX6 and OX62 positive DC and a population devoid of such cells, while S100+ cells were equally divided into these two subpopulations. Although there was a significantly decreased production of IL-6 as compared to that of an original pituitary cell population, both MACS separated populations were equal in IL-6 production. The diminution in IL-6 production in both populations may be the result of an impediment of paracrine communication due to the MACS separation into these two populations. Our data also show that a subpopulation of FS cells was capable of stimulating T cell proliferation in vitro. Concomitantly with the distribution pattern of S100- and OX6-immunoreactive cells in the BSA and Nycodenz gradient fractions, we found a similar pattern of stimulation of T cell proliferation. Unlike the IL-6 production pattern, the T cell stimulating capacity was present in the MHC-class II-enriched cell population but absent in the MHC-class II-depleted cell population. These findings-together with earlier in situ histochemical data-suggest that there is an OX6+ S100- subpopulation of FS cells in the anterior pituitary that in itself is capable of stimulating T cell proliferation in vitro, and acts as lymphoid DC. There is also an S100+ OX6- population that is unable to stimulate T cell proliferation in vitro. Both populations are able to produce IL-6, but probably need stimuli from other subpopulations of pituitary cells (or exogeneous stimuli) to produce maximal amounts of IL-6.

Animals↗