Interleukin 1beta as stimulator of the rat thymus

The effects of interleukin 1beta administration on the thymus of adult and old rats were studied in order to study the interactions between the nervous and immune systems and to confirm the important role played by catecholaminergic nerve fibres (CNF) in the regulation of thymic functions. Moreover, chemical sympathectomy was performed in a group of rats to study the effects on thymus of the destruction of the majority of CNF. Our results indicate that thymic stimulation (performed by means of interleukin 1beta) induces substantial changes in the fresh weight of the whole thymus, as well as in the thymic microenvironment, thymic nerve fibres, CNF, neuropeptide Y (NPY)-like positive nerve fibres and total amount of both proteins and norepinephrine in rat thymic tissue homogenates. The majority of CNF are destroyed after chemical sympathectomy with 6-OH-Dopamine (DA) and remain unchanged after treatment with interleukin 1beta.     

Occurrence of adrenergic nerve fibers in human thymus during immune response

The adrenergic nerve fibers (ANF), the neuropeptide Y-like immunoreactive nerve fibers (NPY-NF) and the noradrenaline (NA) amount were studied in the human thymus in subjects previously treated or not treated with interferon therapy with the aim to identify the changes due to the interferon therapy. This therapy has been used in patients affected by multiple sclerosis (MS). Biochemical and morphological methods were used associated with quantitative analysis of images. The whole thymuses were removed during autopsies in young and adult patients not treated with interferon. Moreover, samples of thymus were removed from patients, either young or adult who had previously been treated with interferon therapy, and subjected, for diagnostic reasons, to thymic biopsy. All samples of thymus were weighed, measured and dissected. Thymic slices were stained with Eosin-orange for detection of the microanatomical details, or with Bodian's reaction for recognition of nervous structures. Histofluorescence microscopy was used for detection of ANF, and immunofluorescence microscopy for recognition of NPY-like immunoreactive structures. All morphological results were subjected to quantitative analysis of images. Noradrenaline contained in thymic structures was measured by biochemical methods. Our results only concerned the effects of the therapy and suggested that treatment with interferon therapy induces many changes in the thymic structures: (1) The protein content of thymus is significantly increased; (2) the NA content in the thymus is also significantly increased; (3) NPY-like immunoreactive structures in the thymus are significantly increased; (4) occurrence of NPY-like immunoreactivity is particularly and significantly increased both in thymic microenvironment and in structures resembling nerve fibers; (5) ANF are significantly increased in the same thymic structures in which NPY-like immunoreactivity is also increased (i.e. thymic microenvironment and structures resembling nerve fibers). The morphological and biochemical changes observed can also explain the immunological changes induced in the thymus after immunostimulating therapy.     

THE INFLUENCE OF ADRENORECEPTOR ACTIVITY ON CRYPT CELL PROLIFERATION IN THE RAT JEJUNUM

The influence of adrenergic stimulation, adrenergic blockade and sympathectomy on crypt cell cycle time and mitotic time in rat jejunum was studied. Alpha adrenergic stimulation by noradrenaline was found to shorten both cell cycle and mitotic times, whereas beta adrenergic stimulation by adrenaline or isoprenaline was found to prolong both cell cycle and mitotic times. Conversely, alpha adrenergic blockade by phentolamine prolonged cell cycle time and beta adrenergic blockade by propranolol or practolol shortened cell cycle time but not mitotic time. Both chemical and surgical sympathectomy inhibited crypt cell proliferation. Chemically sympathectomized animals manifested supersensitivity to exogenous noradrenaline.     

Rat heart GDNF: effect of chemical sympathectomy

Developmental studies indicate a role for GDNF in survival of motor, autonomic, and sensory neurons. However, no study attempted to demonstrate its participation in autonomic nerve regeneration. In this work, chemical sympathectomy by 6-hydroxydopamine provided the model for assessing heart GDNF expression during denervation and axonal regrowth. A glyoxylic acid-based histochemical technique evaluated the noradrenergic innervation. ELISA determined GDNF levels after concentrating heart homogenates. Light and ultrastructural in situ hybridization and immunocytochemistry were used for identifying cells expressing GDNF mRNA and protein. In control rats, the GDNF cardiac levels were significantly higher in 37-day-old animals in comparison with those aging 60 days. In sympathectomized rats, GDNF cardiac levels were significantly higher 7 days after sympathectomy and dropped to control levels at day 30. GDNF mRNA was expressed in atrial and ventricular myocytes from normal and sympathectomized rats. GDNF immunoreactivity occurred on atrial granules and quantitative analysis in electron micrographs confirmed ELISA-obtained data. In ventricular myocytes gold particles occurred sparsely. These findings constitute the first evidence for GDNF synthesis by cardiomyocytes and postulate a role for this factor soon after cardiac sympathetic denervation, probably in nerve regeneration. In atrial myocytes, GDNF is probably secreted by regulated pathway.     

The nerves of the juxtaglomerular apparatus of man and other mammals contain the potent peptide NPY

The juxtaglomerular apparatus, a neuroendocrine unit located in the vascular pole of the glomerulus and influencing blood pressure by the secretion of renin, is known to have a rich supply of monoaminergic nerve fibres. Neuropeptide Tyrosine (NPY), a newly discovered, potent, vasoconstrictor peptide of 36 amino acids, has been found by immunocytochemistry to be present in a dense plexus of fibres around the juxtaglomerular apparatus of man, monkey, mouse, hamster, rat and guinea pig. NPY-immunoreactivity was markedly depleted after chemical sympathectomy by 6-hydroxydopamine. The concentration of NPY within the whole mouse kidney was 29.6 +/- 6.8 pmol/g and fractionation of the extracts demonstrated that the NPY-like immunoreactivity co-eluted from the column in the same position as the porcine NPY standard. The role of this peptide in renal physiology and pathology now needs urgent investigation.     

Influence of surgical and chemical orchidectomy on weight and distribution of AChE-nerve fibres in thymuses of adult rats

The thymus is a crossroad between the immune and neuroendocrine systems. As such, it is innervated by acetylcholinesterase (AChE)-positive fibres of the vagus, the recurrent laryngeal and the phrenic nerves. It is well know, that the innervations density of the thymus increases with age. In our study, adult rats were orchidectomized (surgically and chemically by the application of a luteinizing hormone-releasing hormone). The density of AChE-positive nerve fibres in thymuses, as well as the weight of thymuses was examined. The authors found that both surgical and chemical orchidectomy result in macroscopic and microscopic regeneration of the atrophied thymuses. In regenerated rat's thymuses after orchidectomy the density of AChE-positive nerve fibres was markedly higher in comparison with the control animals. The distribution, as well as the density of AChE-positive nerve fibres in regenerated thymuses after orchidectomy evokes the images of its innervations like in young animals before age-related involution. The authors also found a markedly higher weight of thymuses of orchidectomized rats in comparison with the control groups. In recent study the authors proved that after 8 weeks surgical orchidectomy leads to the regeneration of thymic AChE-positive innervation and chemical orchidectomy by administration of luteinizing hormone-releasing hormone after 4 weeks of adult rats.     

VIP and PHI coexist with an NPY-like peptide in intramural neurones of the small intestine

Vasoactive intestinal peptide (VIP), peptide histidine isoleucine (PHI) and neuropeptide Y (NPY) are neuropeptides present in all layers of the small intestine. NPY-immunoreactive fibres in the gut seem to derive from two sources. One population is of extramural (sympathetic) origin and contains noradrenaline, another is of intramural origin and does not contain noradrenaline. In the present study of mouse, rat and pig, immunocytochemistry showed immunoreactive PHI to coexist completely with immunoreactive VIP. This was predictable, since VIP and PHI derive from the same precursor. In addition, however, VIP and PHI were found to coexist with immunoreactive NPY in non-adrenergic (but not in adrenergic) nerve fibres and nerve cell bodies. This coexistence was unexpected, since the VIP precursor does not contain NPY-like sequences.     

The nerves of the juxtaglomerular apparatus of man and other mammals contain the potent peptide NPY

Summary The juxtaglomerular apparatus, a neuroendocrine unit located in the vascular pole of the glomerulus and influencing blood pressure by the secretion of renin, is known to have a rich supply of monoaminergic nerve fibres.Neuropeptide Tyrosine (NPY), a newly discovered, potent, vasoconstrictor peptide of 36 amino acids, has been found by immunocytochemistry to be present in a dense plexus of fibres around the juxtaglomerular apparatus of man, monkey, mouse, hamster, rat and guinea pig. NPY-immunoreactivity was markedly depleted after chemical sympathectomy by 6-hydroxydopamine. The concentration of NPY within the whole mouse kidney was 29.6±6.8 pmol/g and fractionation of the extracts demonstrated that the NPY-like immunoreactivity co-eluted from the column in the same position as the porcine NPY standard. The role of this peptide in renal physiology and pathology now needs urgent investigation.     

Ethanol-induced cardiac hypertrophy: effects of peripheral sympathectomy

The development of cardiac hypertrophy was examined in rats that had undergone sympathectomy with 6-hydroxydopamine. After 4 days, the rats were given severely intoxicating doses of ethanol or isocalorically paired quantities of maltose-dextrin by intubation at 8-h intervals up to 48 h. The ethanol and sugar intubations were applied in a nutritionally adequate, liquid diet mix. The extent of the peripheral sympathectomy was evident from the absence of detectable quantities of noradrenaline in hearts of animals injected with the neurotoxin and in the reduced levels of excreted noradrenaline. The adrenal medullary catecholamine contents of sympathectomized rats were unchanged in the absence of ethanol; there were reduced quantities of adrenaline in the medullae of rats given ethanol. The adrenal glands of rats given ethanol were larger than those from control animals. Urine samples from sympathectomized and control rats, given ethanol, displayed equivalent increases in excreted adrenaline and noradrenaline. Increases in relative cardiac weight were evident in hearts from sympathectomized animals after 4 days of sympathectomy, and this change reached significance in the hearts from 6-hydroxydopamine-treated rats after a further 2 days on the control diet. Hearts from animals exposed to ethanol showed a marked, rapid development of cardiomegaly; after 24 h there was an increased mass of some 17%, which was sustained over the remaining 24-h period. The proportion of cardiac protein did not differ in the large hearts from ethanol-treated animals and those from their controls, hence myocardial oedema could not account for the increase in weight.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immuno-characterisation of neuroendocrine cells of the rat thymus gland in vitro and in vivo

Primary cell cultures and organ fragments of rat thymus were characterised by use of a panel of antibodies raised against the neural adhesion molecule L1, tyrosine hydroxylase, protein gene product 9.5, nerve growth factor, calcitonin gene-related peptide, glial fibrillary acidic protein, vimentin, pan-cytokeratin, a ganglioside of neural crest and neuroendocrine cells (A2B5), and thymulin (4 beta). Immunoreactivity for neural markers only was identified in a single morphology (nerve-like) of cell in culture and throughout the adult thymus as fine, tortuous staining. Immunoreactivity for endocrine markers only was identified in polygonal epithelial-like cells in culture, throughout viable tissue in fragment culture and in the subcapsular cortex of the adult thymus. Immunoreactivity for both endocrine and neural markers was identified in three distinct morphologies in cell culture: elongate, spherical, and stellate. Similar results were observed in the mitotic periphery of the cultured fragments and in the medulla and cortico-medullary junction of the adult thymus. Cells with immunoreactivity to A2B5 were present in primary and fragment cultures and occurred throughout the adult thymus. The apparent diversity of cell immunoreactivity in primary and fragment thymic cultures suggests that numerous neural and endocrine factors may be required for the development and/or regeneration of the thymic microenvironment.     

Quantitation and immunohistochemistry of catecholamines in the posterior segment of the eye

Summary Dopamine, noradrenaline, and adrenaline were assayed with HPLC in the light adapted retinae of carp, frog, chicken, pigeon, rat, guinea-pig, rabbit, cat, pig and cow. Dopamine varied from 0.6 to 2.6 nmol/g wet weight and was not influenced by sympathectomy. The dopamine figures agree with previously published results. Noradrenaline concentrations varied from not detectable to 0.06 nmol/g wet weight in different species. Homolateral sympathectomy significantly decreased the noradrenaline figure in rabbits. There are no previous figures for noradrenaline for most of the species. Adrenaline was not detected in any species. Immunohistochemical analysis showed noradrenaline to be present in choroidal nerves, but noradrenaline immuno-reactivity was not seen in the retina (chicken, rat, guinea-pig, rabbit, cat, cow). It is concluded that dopamine is the major catecholamine in the retina. Noradrenaline was found present only in minute amounts in the assays, and much of its was likely to stem from sympathetic nerve fibres. The study did not demonstrate any noradrenergic neurons in the retina.     

Chemical sympathectomy with 6-hydroxydopamine potentiates intracerebroventricularly applied bombesin-induced increase in plasma adrenaline

Intracerebroventricular administration of bombesin induced a marked increase in plasma level of adrenaline and a slight increase in that of noradrenaline in rats anesthetized with urethane. The bombesin-induced increase in adrenaline was potentiated by chemical sympathectomy with 6-hydroxydopamine (6-OHDA). On the other hand, adrenalectomy did not affect plasma level of noradrenaline in the bombesin-treated animals. In the splanchnicotomized rats, direct stimulation of the adrenal glands by intravenously administered nicotine increased plasma level of both adrenaline and noradrenaline. These increases were, however, not potentiated by chemical sympathectomy with 6-OHDA. Pretreatment with capsaicin, a potent toxin selective to sensory neurons, potentiated the bombesin-induced increase in plasma level of adrenaline. In these capsaicin pretreated rats, chemical sympathectomy did not potentiate the bombesin-induced increase in plasma level of adrenaline to any great extent. These results suggest that chemical sympathectomy with 6-OHDA potentiated the bombesin-induced increase in plasma adrenaline probably due to a disinhibitory activation of the splanchnic nerve by as yet unidentified but capsaicin sensitive neuron mechanisms.     

Age changes in rat vasomotor reflexes and sympathetic neuron ultrastructure following chemical sympathectomy]

Prolonged administration of guanethidine (20 mg/kg) to newborn rats caused a marked reduction in the number of cells in stellate ganglia. The administration of guanethidine for 14 days decreased the amount of cells to 30% of the normal (partial sympathectomy), and for 28 days--to 0.5% (complete sympathectomy). At the age of two months the blood pressure pressor reflexes to asphyxia and femoral nerve stimulation were absent in both groups of the sympathectomized animals. These responses, however, were restored in the partially sympathectomized animals at the age of four months. No restoration took place in the completely sympathectomized animals. The electron microscopic studies of neurons in the partially sympathectomized animals showed the presence of a great number of neurofibrils. According to literature data this fact was typical of cells in which an active growth of axon fibers took place.     

Chemical sympathectomy and diurnal fluctuations of noradrenaline calorigenic action in the rat

In the rat, in which a diurnal fluctuation of the sensitivity to noradrenaline was previously found, the effect of injected 6-hydroxydopamine (6-OHDA) was investigated. The heat production and catecholamines contents in the interscapular brown adipose tissue, heart and adrenals were measured. Chemical sympathectomy induces a disappearance of diurnal fluctuation in the sensitivity to injected noradrenaline. In these animals a lower capacity for heat production was found. However, a significant calorigenic effect of injected noradrenaline in 6-OHDA-treated animals was still present. In sympathectomized animals a depletion of noradrenaline from interscapular brown adipose tissue and the heart was observed. Besides, a change in adrenaline/noradrenaline ratio was found in the adrenals.     

Neuropeptide Y-like immunoreactivity in intramural ganglia of the newborn guinea pig urinary bladder

Immunoreactive neuropeptide Y (NPY) was demonstrated in neuronal elements in the urinary bladder wall of the newborn guinea pig. Numerous intramural ganglia were found lying among the smooth muscle bundles and in the submucosa, and NPY-like immunoreactive nerve cell bodies were demonstrated within all of these ganglia. Nerve fibres containing NPY were also richly distributed in the detrusor muscle, submucosa and around blood vessels. In dissociated cell cultures from newborn guinea pig detrusor muscle, a subpopulation (70-85%) of both mononucleate and binucleate intramural neurones was shown to contain NPY-like immunoreactivity. A low percentage (1-6%) of the intramural bladder neurones contained dopamine-beta-hydroxylase. In conclusion, while some NPY-containing nerve fibres in the wall of the bladder are of sympathetic origin, especially those supplying blood vessels, the results of this present study establish that many of these NPY-containing nerve fibres originate from non-adrenergic cell bodies within the intramural bladder ganglia.     

Immunosenescence: potential causes and strategies for reversal

Age-related deterioration in immune function has been recognized in many species. In humans the clinical manifestation of such immune dysfunction is age-related increases in the susceptibility to certain infections and in the incidence of some autoimmune disease and certain cancers. Laboratory investigations reveal age-related changes in the peripheral T cell pool, in the predominant phenotype, cytokine production profiles, signalling function and in replicative ability following stimulus with antigen, mitogens or anti-CD3 antibody. These changes in the properties of peripheral T cells are thought to be causally linked to an age-associated involution in the thymus. Our analysis reveals that thymic involution is due to a change in the thymic microenvironment linked to a reduction in the level of available interleukin 7. Treatment with interleukin 7 leads to a reversal of thymic atrophy with increased thymopoiesis. This provides the potential to reverse the immune dysfunction seen in the peripheral T cell pool by replacing old cells with new output generated in the thymus. Problems to overcome in order for such an experimental therapy to be successful require careful analysis in order to provide an optimal strategy to ensure that new T cell emigrants from the thymus have a broad range of specificities and are able to enter the peripheral T cell pool.     

Effects of pregnancy on the extrinsic innervation of the guinea pig uterus. A histochemical, immunohistochemical and ultrastructural study

Summary The extrinsic innervation of the guinea pig uterus was studied by immunohistochemical, ultrastructural and enzyme histochemical methods.The extrinsic innervation was organized in two major ways. One consisted of nerve trunks and non-varicose nerve fibres running in the suspensory ligament, and the other of a plexus of varicose nerve fibres surrounding vessels, and non-vessel-related non-varicose nerve fibres in the mesouterus. The use of different neuronal and Schwann cell markers showed that the extrinsic innervation was predominantly adrenergic and contained only few peptidergic nerves. Acetylcholinesterase-positive (cholinergic) nerves were only found around the uterine artery.In late pregnancy, the extrinsic nerves of the mesouterus adjacent to foetus-containing uterine horns underwent pronounced degenerative changes comprising both Schwann cell and axonal structures. In comparison, no changes were found in extrinsic nerves of mesouteri adjacent to non-foetus-bearing uterine horns or in extrinsic nerves in the suspensory ligaments. Further, chemical sympathectomy produced axonal degeneration but no changes in the Schwann cells.In conclusion, the pregnancy-induced nerve degeneration is of a very special type different from that following chemical sympathectomy and represents a local phenomenon related to the conceptus. Hypothetically, this could be of importance for counteracting disturbances in placental blood flow.     

The thymus microenvironment in regulating thymocyte differentiation

The thymus plays a crucial role in the development of T lymphocytes by providing an inductive microenvironment in which committed progenitors undergo proliferation, T-cell receptor gene rearrangements and thymocyte differentiate into mature T cells. The thymus microenvironment forms a complex network of interaction that comprises non lymphoid cells (e.g., thymic epithelial cells, TEC), cytokines, chemokines, extracellular matrix elements (ECM), matrix metalloproteinases and other soluble proteins. The thymic epithelial meshwork is the major component of the thymic microenvironment, both morphologically and phenotypically limiting heterogeneous regions in thymic lobules and fulfilling an important role during specific stages of T-cell maturation. The process starts when bone marrow-derived lymphocyte precursors arrive at the outer cortical region of the thymic gland and begin to mature into functional T lymphocytes that will finally exit the thymus and populate the peripheral lymphoid organs. During their journey inside the thymus, thymocytes must interact with stromal cells (and their soluble products) and extracellular matrix proteins to receive appropriate signals for survival, proliferation and differentiation. The crucial components of the thymus microenvironment, and their complex interactions during the T-cell maturation process are summarized here with the objective of contributing to a better understanding of the function of the thymus, as well as assisting in the search for new therapeutic approaches to improve the immune response in various pathological conditions.Key words: thymus, T-cell maturation, thymic microenvironment, thymocyte differantiation, chemokines, extracellular matrix, thymic nurse cells, metalloproteinases     

The thymus microenvironment in regulating thymocyte differentiation

The thymus plays a crucial role in the development of T lymphocytes providing an inductive microenvironment in which committed progenitors undergo proliferation, T-cell receptor gene rearrangements and thymocyte differentiation into mature T-cells. The thymus microenvironment forms a complex network of interaction that comprises non lymphoid cells (e.g., thymic epithelial cells, TEC), cytokines, chemokines, extracellular matrix elements (ECM), matrix metalloproteinases and other soluble proteins. The thymic epithelial meshwork is the major component of thymic microenvironment, both morphologically and phenotypically limiting heterogeneous regions in thymic lobules and fulfilling an important role during specific stages of T-cell maturation. The process starts when bone marrow–derived lymphocyte precursors arrive at the outer cortical region of the thymic gland and begin to mature into functional T lymphocytes that will finally exit the thymus and populate the peripheral lymphoid organs. During their journey inside the thymus, thymocytes must interact with stromal cells (and their soluble products) and extracellular matrix proteins to receive appropriate signals for survival, proliferation and differentiation. The crucial components of the thymus microenvironment and their complex interactions during the T-cell maturation process with the objective of contributing to a better understanding of the function of the thymus as well as assist in the search for new therapeutic approaches to improve the immune response in various pathological conditions are summarized here.     

Occurrence and distribution of neuropeptide-Y-immunoreactive nerves in the respiratory tract and middle ear

Summary Nerve fibres displaying neuropeptide-Y (NPY) immunoreactivity are abundantly distributed in the respiratory tract of cats, guinea-pigs, rats and mice. Fine beaded NPY fibres were seen in whole-mount spreads of the middle-ear mucosa. In the nasal mucosa and in the wall of the Eustachian tube NPY fibres were numerous around arteries and arterioles but sparse in the vicinity of veins; single fibres were found close to the acini of seromucous glands. In the tracheobronchial wall NPY fibres occurred in the proximity of blood vessels, in the subepithelial layer and in the smooth muscle. Surgical and chemical (6-hydroxydopamine treatment) sympathectomy resulted in disappearance of adrenergic and NPY-containing nerve fibres in the nasal mucosa. Sequential staining with antibodies against dopamine--hydroxylase (DBH) and NPY revealed that DBH and NPY occur in the same perivascular nerve fibres in the nasal mucosa. The distribution of NPY fibres in the respiratory tract suggests multiple functions of NPY, such as regulation of local blood flow, glandular secretion and smooth muscle activity.