Cytokines as modulators of the hypothalamus-pituitary-adrenal axis.

The hypothalamus-pituitary-adrenal (HPA) axis is stimulated during the course of certain immune, inflammatory and neoplastic processes. IL-1 is an important immunologically derived cytokine mediating the stimulation of this axis, although not the only one. We have compared the relative potencies of the cytokines IL-1, IL-6 and tumor necrosis factor (TNF), which share several biological actions, for stimulating ACTH and corticosterone output in freely-moving rats. Although all three cytokines can stimulate the HPA axis, IL-1 was the most potent. This effect of IL-1 was also present during the neonatal period, when the response of the HPA axis to acute stress is reduced in rodents. The results support the existence of an immune-HPA axis circuit. The biological and clinical relevance of this circuit is discussed.

Administration of interleukin-1 at birth affects dopaminergic neurons in adult mice.

Interleukin-1 beta (IL-1 beta), a cytokine that plays a relevant role during inflammatory and immune processes, can also affect brain neurotransmitters and the activity of peripheral sympathetic nerves. Because both brain and peripheral catecholaminergic systems in mice are not fully developed at birth, we speculated that the development of these systems may be susceptible to modifications when mice are exposed to IL-1 beta early in life. Here we report that the administration to mice of a low dose of IL-1 beta during the first days of life results in a decreased dopamine content in the hypothalamus in adulthood. We also show that the dopamine content of the superior cervical sympathetic ganglia was reduced in adult mice that were treated with IL-1 beta at birth. No changes in noradrenaline content nor in its metabolite MHPG were detected in the brain and peripheral sympathetic ganglia of these animals. This indicates that central and probably also peripheral dopaminergic neurons are preferentially affected by IL-1 beta treatment at birth. Collectively, these results indicate that an increased production of IL-1 beta during infectious or inflammatory processes in the perinatal period may induce long-lasting, probably permanent, alterations in central and peripheral neurotransmitter systems.

Sympathetic innervation affects superantigen-induced decrease in CD4V beta 8 cells in the spleen.

The stimulation by superantigens of T cells expressing an appropriate V beta chain results in a strong proliferative response that is followed by a state of energy specific for the antigen used. This model was used to continue our studies on immunoregulatory host neuroendocrine responses. We have recently found that four days after administration of the superantigen staphylococcal enterotoxin B (SEB) into mice, that is, at an early stage of the anergic phase, the decrease in the percentage of splenic CD4V beta 8 was accompanied by a decrease in the splenic concentration of the sympathetic neurotransmitter noradrenaline (NA) as compared to vehicle-injected mice. No comparable changes were detected in the kidney. At this point, blood levels of NA, adrenaline, and corticosterone were comparable in SEB- and vehicle-injected mice. We have also found that the decrease in splenic CD4V beta 8 cells was not observed in animals that had been chemically sympathectomized prior to the administration of the superantigen. These results indicate that the sympathetic response induced by SEB may have immunoregulatory implications.

Monoamines and related substances in brainstem and spinal cord of Lewis rats during the attack and recovery of experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis influenced catecholaminergic and indoleaminergic neurotransmitter systems in the central nervous system of Lewis rats. During paralysis, serotonin and noradrenaline were significantly reduced compared to animals injected with complete Freund's adjuvant in the posterior dorsomedial brainstem and in lower spinal cord segments. These diminutions remained after recovery from neurological signs in T11-S1. The serotonin metabolite 5-hydroxyindoleacetic acid was greatly augmented during the attack in all segments but was depleted during recovery in the lumbar spinal cord, which may indicate a normalized turnover at a reduced serotonin level. These results suggest functional impairment of monoaminergic neurons of the brainstem and spinal cord followed by permanent damage to some monoaminergic fibers in the spinal cord.

Levels of low molecular weight scavengers in the rat brain during focal ischemia.

Ascorbic acid, cysteine, glutathione, uric acid, tyrosine and tryptophan were quantified in samples of frontoparietal cortex, striatum, hippocampus and cerebellum from both sides of rat brain 0.5 h, 4 h and 24 h after focal ischemia. Cysteine, tyrosine and tryptophan were increased in cortex and striatum at 0.5 h, returning afterwards to normal. Uric acid was increased, whereas ascorbic acid and glutathione were correspondingly decreased. Although changes can be explained primarily by energy failure they are also consistent with free radical activity during early stages of ischemia.

Transformations of the retinal topography along the visual pathway of the chicken.

It is still unclear how the retinotectal map of the chick is formed during development. In particular, it is not yet known whether or not the organization of fibres plays a role in the formation of this map. In order to contribute to the solution of this problem, we analysed the representation of the retinal topography at closely spaced intervals along the fibre pathway. We injected HRP into various sites of the tectal surface and traced the labelled fibre bundles back to the retina. The retinal topography was reconstructed at ten different levels, i.e. in the retina, the optic nerve head, the middle of the optic nerve, the chiasm (three levels), the optic tract (three levels), and the optic tectum. We obtained the following results: (1) The labelled fibre bundles as well as the fields of labelled retinal ganglion cells were always well delimited and coherent. (2) The reconstructions show that transformations of the retinal topography occur in the fibre pathway. The first and most important transformation is found in the optic nerve head where the retinal image is mirrored across an axis extending from dorsotemporal to ventronasal retina. In addition, the retinal representation is split in its temporal periphery. Thus, central and centrotemporal fibres are no longer in the centre of the image but close to the dorsal border of the nerve. Peripheral fibres are found along the medial, ventral and lateral circumference of the nerve. In the optic tract a second transformation occurs. The retinal topography is rotated clockwise by about 90 degrees and flattened to a band. The flattening is accompanied by a segregation of fibre bundles so that eventually central and centrotemporal retinal fibres are located centrally, ventral fibres dorsally and dorsal retinal fibres ventrally in the tract. By these two transformations an organization of fibres is produced in the optic tract which can be projected onto the tectal surface without major changes given that dorsal and ventral fibres remain in their relative positions, and that deep lying fibres project to the rostral and central tectum, superficial fibres to the caudal tectum. The transformations which we have observed follow specific rules and thus maintain order in the pathway although retinotopy is lost. In conjunction with our earlier studies on the development of the retinotectal system we conclude that fibres are laid down in a chronotopic order. The transformations take place under particular structural constraints.(ABSTRACT TRUNCATED AT 400 WORDS)

Measurement of low-molecular-weight antioxidants, uric acid, tyrosine and tryptophan in plaques and white matter from patients with multiple sclerosis.

The levels of the antioxidants ascorbic acid, cysteine, reduced glutathione and alpha-tocopherol, of the free-radical marker uric acid and of the amino acids tyrosine and tryptophan were measured by means of high-pressure liquid chromatography in plaques, adjacent white matter and distant white matter from patients with multiple sclerosis, and in central nervous system tissue from patients without neurological diseases. Cholesterol and DNA were also determined, to check demyelination and cellularity. Uric acid was increased and glutathione correspondingly decreased in plaques; alpha-tocopherol was lowest in plaques and highest in distant white matter in all cases. Ascorbic acid, cysteine, tyrosine and tryptophan were not significantly changed in any tissue. The results provide evidence supporting the involvement of free radicals in multiple sclerosis.

Induction of cytokine transcripts in the central nervous system and pituitary following peripheral administration of endotoxin to mice.

The regional distribution and inducibility of cytokines in the normal brain is still a matter of controversy. As an attempt to clarify this issue, we studied the constitutive and induced expression of interleukin (IL)-1beta, IL-6, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma mRNAs in the brain, pituitary, and spleen of mice using qualitative and semiquantitative reverse-transcription polymerase chain reaction. The contribution of nonbrain cells to the cytokine transcripts detected was considered. With the exception of IFN-gamma mRNA, transcripts for the other cytokines were found to be constitutively present in the brain. Following i.p. injection of lipopolysaccharide (LPS) at a dose below those described to disrupt the blood-brain barrier (BBB), cytokine mRNA expression was increased in the spleen, the pituitary, and the brain. In the brain, the onset of transcription varied from 45 min (IL-1beta, TNF-alpha) to 4 hr (IFN-gamma), and the peak of mRNA accumulation was observed at different times depending on the cytokine and the brain region studied. IL-1 and IL-6 were highly expressed in the hypothalamus and hippocampus, while TNF-alpha expression was more marked in the thalamus-striatum. The cortex was the region in which cytokines were less inducible. The inducible expression of cytokine mRNAs in the brain was paralleled by stimulation of the hypothalamus-pituitary-adrenal axis. These results show the capacity of brain cells to synthesize different cytokine mRNAs in vivo and define the kinetics of their expression in several brain areas and in the periphery in parallel to the activation of a neuroendocrine pathway by endotoxin.

Quantitative determination of water-soluble scavengers in neoplastic and non-neoplastic human breast tissue.

The water-soluble scavengers ascorbic acid (Asc), cysteine (Cys), glutathione (GSH) and uric acid (UA) as well as DNA content were determined in 40 breast tissue samples (neoplastic and non-neoplastic tissues from 20 patients). To allow proper homogenization to take place, a fixed number of sections was cut from a tissue cylinder of known diameter. Adjacent sections were stained with hematoxylin-eosin and the fractions of epithelium, fat and connective tissue were estimated as a percentage of the section area. Protein-free extracts were injected into a reverse-phase high-pressure liquid chromatography system and scavengers quantified with 2 electrochemical detectors (gold and glassy carbon). DNA and all scavengers, except UA, were greatly increased in cancer tissues in nearly all cases. Amounts of Asc and GSH in neoplastic tissue correlated closely with DNA values and percentage of epithelium, those of Cys not so closely and those of UA not at all. We assume that Asc and GSH were located mainly in the epithelium, UA mainly in the extracellular space and Cys in both spaces. When values were expressed as mumol/g DNA, a parameter related to content per cell, values were higher in neoplastic than in non-neoplastic tissue for Asc (18/20 cases), GSH (17/20) and Cys (14/20) and lower in neoplastic tissue for UA (19/20). It is known that increased GSH protects cells against certain drugs in tissue cultures. For in vivo treatment the presence of increased Asc (and to a lesser extent Cys) in addition to GSH could be of importance.

Endotoxin impedes vasoconstriction in the spleen: role of endogenous interleukin-1 and sympathetic innervation.

Processes relevant for an appropriate immune response such as immune cell traffic and recirculation require a tight control of blood supply to lymphoid organs. Interactions between endogenous cytokines and sympathetic nerve fibers in lymphoid organs can contribute to this control. The results reported in this paper show that 1) administration of low doses of lipopolysaccharide (LPS), an endotoxin derived from gram-negative bacteria, causes an increase in splenic blood flow (SBF); 2) this increase is mediated by the production of endogenous interleukin-1 (IL-1); 3) the effect of LPS on SBF requires an intact splenic sympathetic innervation; 4) the LPS-induced increase in SBF is exerted at the postganglionic level; 5) the endotoxin inhibits the vasoconstriction induced by the in vivo stimulation of the splenic nerve but does not affect the vasoconstriction induced by norepinephrine (NE); and 6) although IL-1 and LPS stimulate general sympathetic activity as reflected by increased peripheral vascular resistance, they do not increase NE concentration in splenic dialysates. Together these in vivo results indicate that endogenous IL-1 affects blood supply to the spleen by inhibiting the sympathetic vasoconstrictor tonus at a postganglionic, prejunctional level. This effect is expected to be relevant for immune cell recirculation, homing, and traffic as well as antigen trapping in the spleen, an organ specialized in the control of these processes during immune responses.

Quantitative determination of water- and lipid-soluble antioxidants in neoplastic and non-neoplastic human breast tissue.

Ascorbic acid, cysteine, glutathione and uric acid were determined by reverse-phase high-pressure liquid chromatography (HPLC) in 46 breast tissue samples [neoplastic (C) and non-neoplastic (N) from the same patient]. Cholesterol, alpha-tocopherol and gamma-tocopherol were quantified in 64 similar samples by extraction into heptane followed by direct-phase HPLC. DNA was measured in all samples and the percentages of epithelium, fat and connective tissue were estimated in sections adjacent to the sample. Results confirm previous findings that ascorbic acid and glutathione, expressed as mumol/g DNA, were greatly increased in the epithelium of neoplastic tissue. Similar increases in cysteine could be accounted for by the presence of inflammatory cells. Although values of alpha- and gamma-tocopherol correlated with the percentage of fat in both types of tissue, these compounds were also present in the epithelium. Because of the varying amounts of fat in the samples, no significant difference could be found between N and C values. Cholesterol correlated with fat in N and epithelium in C. Consideration of 10 cases with equal amounts of fat in C and N tissue suggests that cholesterol is reduced in C in the epithelial cells.

Interleukin-1 induces changes in norepinephrine metabolism in the rat brain.

Interleukin-1 (IL-1) is a hormone that, apart from playing a key role in immune and inflammatory processes, can also affect mechanisms under brain control. To gain a better understanding of the action of this cytokine on the CNS, its effects on the contents of norepinephrine (NE), dopamine (DA) and serotonin (5-HT), and their main metabolites and precursors, were evaluated in different regions of the forebrain, brain stem, and spinal cord. Following administration of human recombinant IL-1 (beta form) to rats, a modest decrease in the content of NE was observed in the hypothalamus as well as in the dorsal posterior brain stem. However, the most relevant finding was that 3-methoxy-4-hydroxyphenylethylene glycol (MHPG), the main NE metabolite, and the relation MHPG/NE were increased in all the regions studied, revealing a stimulatory effect of IL-1 on NE metabolism in the CNS. This effect seems to be specific for NE since no comparable changes in the brain content of DA, 5-HT, or its metabolite, 5-hydroxyindole acetic acid, were detected after administration of the cytokine. However, tryptophan was significantly increased in all brain regions and in the cervical spinal cord. The capacity of IL-1 to affect the metabolism of NE, a neurotransmitter involved in the control of a variety of brain functions, provides further proof for the relevance of this cytokine in brain-immune interactions.

Interleukin-1 increases splenic blood flow by affecting the sympathetic vasoconstrictor tonus.

The possibility that interleukin-1 (IL-1), a cytokine involved in immune and inflammatory mechanisms, can affect the blood flow of the spleen was considered because changes in spleen perfusion can affect immune cell recirculation, traffic, and homing. The results indicate that administration of a subpyrogenic dose of IL-1 induced a pronounced increase in splenic blood flow. This was not a general effect, because no change in blood flow of skeletal muscle was noticed. The studies also show that 1) the increase in splenic perfusion induced by IL-1 is to a large extent independent from the secondary induction of nitric oxide (NO), 2) the splenic blood flow in the rat is under sympathetic control, and 3) the effect of IL-1 on splenic blood flow is completely abrogated after surgical interruption of the splenic nerve, which is predominantly composed of sympathetic fibers. It is concluded that the IL-1-mediated increase in splenic blood flow is most likely based on the inhibition of the sympathetic vasoconstrictor tonus in the rat spleen. These results show that a cytokine released by activated immune cells can regulate the blood flow of a main lymphoid organ, the spleen, by affecting mechanisms under neural control.

Alterations in the pituitary-adrenal axis of adult mice following neonatal exposure to interleukin-1.

Interleukin-1 (IL-1), a cytokine mainly derived from activated cells of the macrophage lineage, can stimulate the hypothalamus-pituitary-adrenal (HPA) axis. Acute and long-lasting effects on the HPA axis were induced by the administration of low doses of IL-1 to mice during the first 5 days of life. In 5-day-old mice, corticosterone blood levels were markedly elevated 2 h following the last injection of IL-1. IL-1-treated mice grew normally. When studied during adulthood, however, these animals showed a reduction in morning values of corticosterone and the ACTH/corticosterone ratio was increased. Furthermore, an inverse correlation between ACTH and corticosterone levels in blood and between ACTH content in the pituitary gland and corticosterone levels was observed in IL-1-treated mice. Lower blood levels of corticosterone were not due to a reduced sensitivity of the adrenal glands, because these animals responded normally to exogenous ACTH. Another alteration observed in IL-1-exposed adult mice was a reduction in ACTH-like immunoreactivity in the pituitary gland following acute cold and restraint stress. It is concluded that exposure of mice to IL-1 early in life causes long-lasting alterations in the HPA axis. Spleen cells from adult mice treated with IL-1 at birth also developed a stronger response to allogeneic antigens than did cells from control mice. This observation indicates the relevance of immune-neuroendocrine interactions during development.