Inducible expression of the long pentraxin PTX3 in the central nervous system.

PTX3 is a prototypic long pentraxin consisting of a C terminal 203-amino acid pentraxin-like domain coupled with an N-terminal 178-amino acid unrelated portion. PTX3 is induced by primary proinflammatory signals in various cell types, most prominently macrophages and endothelial cells. Other long pentraxins, such as murine or rat neuronal pentraxin 1 (NP1) and human neuronal pentraxin 2 (NPTX2), are expressed in the central nervous system (CNS). The present study was designed to investigate whether PTX3 is expressed in the brain and to define the structures and cells involved. Intracerebroventricular (i.c.v.), but not i.v., injection of LPS induced high levels of PTX3 mRNA in the mouse brain. In contrast NP1 is constitutively expressed in the murine CNS and is not modulated by LPS administration. I.c.v. IL-1beta was also a potent inducer of PTX3 expression in the CNS, whereas TNFalpha was substantially less effective and IL-6 induced a barely detectable signal. Central administration of LPS and IL-1 induced PTX3 also in the periphery (heart), whereas the reverse did not occur. Expression of PTX3 was also observed in the brain of mice infected with Candida albicans (C. albicans) or Cryptococcus neoformans. (C. neoformans). The kinetics of PTX3 gene induction were consistently different between C. albicans- and C. neoformans-infected mice, according to the diverse outcome of the CNS immune reaction. In situ hybridization revealed that i.c.v. injection of LPS induced a strong PTX3 expression in presumptive glial cells, in the white matter (corpus callosum, fimbria) and meningeal pia mater as well as in dentate gyrus hilus and granule cells. No constitutive expression of PTX3 was detected. Central expression of PTX3 may amplify mechanisms of innate resistance and damage in the CNS. The possibility of a direct interaction of PTX3 with neuronal cells, as suggested for NPTX2, remains to be explored.

Increased peripheral benzodiazepine binding sites and pentraxin 3 expression in the spinal cord during EAE: relation to inflammatory cytokines and modulation by dexamethasone and rolipram.

We have studied the mRNA expression of pentraxin 3 (PTX3) and the binding of the peripheral-type benzodiazepine receptor (PBR) ligand, [3H]-PK11195, in the spinal cord of Lewis rats where EAE was actively induced. PTX3 was induced during the active phase of EAE (day 10-14), it remained high up to 30 days and disappeared only 60 days later. Similarly, PK11195 binding peaked at day 14-17 during the recovery and it disappeared by day 60. On the other hand, the levels of TNF and IL-6 in the spinal cord were elevated at the peak and at the onset of clinical signs and returned to non-detectable by day 14-17. Dexamethasone abolished all these changes, while treatment with rolipram, delayed the appearance of the disease and then decreased its severity. However the peaks of TNF, IL-6, PBR and PTX3 levels in spinal cord were only delayed, but not reduced, by rolipram treatment. In conclusion, we show two types of inflammatory changes in EAE: acute, short term changes (TNF and IL-6), that correlate with the disease; and effects such as PTX3 expression and PK11195 binding that last longer after recovery from the disease.

Increased tumor necrosis factor and interleukin-6 production in the central nervous system of interleukin-10-deficient mice.

Interleukin-10 (IL-10) inhibits tumor necrosis factor (TNF) production. We investigated the role of endogenous IL-10 in brain TNF production. We injected IL-10-knockout mice with lipopolysaccharide (LPS,2.5 microg/mouse i.c.v.). Brain TNF and IL-6 levels were more elevated and persisted longer in IL-10-deficient mice compared with wild type mice, suggesting that IL-10 is an important negative feedback inhibitor of TNF and IL-6 production in the CNS.

Childhood obesity: results of a multicenter study of obesity treatment in Italy.

The prevalence of pediatric obesity is increasing and many patients are followed by specialized centers or private doctors. The aim of this study was to verify short- and medium term results of a therapeutic approach based on nutritional intervention in a large pediatric population: 1383 subjects (695 females, 688 males) aged 10.1 +/- 2.7 yr, followed in 11 pediatric departments in Italy. No difference was found between centers in age, height, weight, BMI and IBW. The drop-out rate after the first visit was 30.2% (58.1% IBW > 140%) in females and 34.2% (70.7% IBW > 140%) in males. After two years of follow-up only 9.7% of females and 6.4% of males remained on treatment. Of these patients only 7.3% of females and 6.4% of males had IBW < 120%. These data show that an approach based on nutritional intervention alone is not sufficient for long-term treatment of pediatric obesity. Only an approach started early and involving the family can produce permanent results.

Decrease in brain cytochrome P450 enzyme activities during infection and inflammation of the central nervous system.

The effect of infection and inflammation of the central nervous system (CNS) on cytochrome-P450-dependent activities in brain, spinal cord and liver microsomes was determined. For this, two models were used: (1) the intracerebroventricularly injected lipopolysaccharide (LPS) model and (2) the experimental auto-immune encephalomyelitis (EAE) model. In the LPS model, aminopyrine N-demethylase (AMND) and ethoxycoumarin O-deethylase (ECOD) activities (both P450 dependent) were significantly decreased (35 and 20%, respectively) in brain microsomes. In the EAE model, only ECOD activity was significantly lower (18%). In the liver, a decrease in total P450, AMND and ECOD activities was only observed in the LPS model. In both models, tumour necrosis factor (TNF) was significantly elevated in brain and spinal cord tissues. In serum, TNF was only detectable in the LPS model. It is concluded that an infection or inflammation located in the CNS, which is accompanied by high TNF levels, results in a decrease in P450-dependent metabolism not only in the liver but in the brain as well.

Lps induces IL-6 in the brain and in serum largely through TNF production.

We investigated the relative contribution of IL-6 and PGE2 directly induced by LPS and indirectly induced via TNF, using in vivo and in vitro models in the mouse. In these models we have used as tools an anti-TNF antibody and a cyclooxygenase inhibitor, the S enantiomer of ketoprofen (S-KPF). Anti-TNF antibodies inhibited LPS-induced IL-6 production in three different models: IL-6 production by mouse peritoneal macrophages in vitro; serum IL-6 levels induced by intraperitoneal LPS; and brain IL-6 levels induced by an intracerebroventricular injection of LPS. However, in vitro anti-TNF antibodies, did not inhibit LPS-induced PGE2, indicating that this effect is not mediated by TNF. Since PGE2 has an opposite effect on TNF and IL-6 production, inhibiting that of TNF but inducing that of IL-6, we investigated the effect of S-KPF on TNF and IL-6 production in vivo following LPS injection. Both TNF and IL-6 induction was augmented by S-KPF, but anti-TNF antibodies abolished the augmentation of IL-6 production. Thus, the effect of anti-inflammatory drugs on IL-6 production in some models can be secondary to their effect on TNF production.

Nonsteroidal anti-inflammatory drugs increase tumor necrosis factor production in the periphery but not in the central nervous system in mice and rats.

Nonsteroidal anti-inflammatory drugs (NSAIDs), which inhibit prostaglandin (PG) synthesis, augment production of tumor necrosis factor (TNF) in most experimental models. We investigated the effect of two NSAIDs, indomethacin and ibuprofen, on the production of TNF in the CNS induced by intracerebroventricular injection of lipopolysaccharide (LPS). Indomethacin and ibuprofen, administered intraperitoneally, augmented (three- to ninefold) the levels of TNF in serum and peripheral organs of mice injected intraperitoneally with LPS and in rats with adjuvant arthritis (up to a sevenfold increase). However, NSAIDs (intraperitoneally or intracerebroventricularly) did not increase brain TNF production induced by intravenous LPS. In fact, indomethacin decreased (1.4-1.8-fold) TNF levels in the spinal cord of rats with experimental autoimmune encephalomyelitis and in the cortex of rats with focal cerebral ischemia. Systemic administration of iloprost inhibited serum TNF levels after intraperitoneal LPS, whereas intracerebroventricular injection of iloprost or PGE2 did not inhibit brain TNF induced by intracerebroventricular LPS. Both peripheral and central TNF productions were inhibited by cyclic AMP level-elevating agents or dexamethasone. Thus, a PG-driven negative feedback controls TNF production in the periphery but not in the CNS.

Corticosteroid-independent inhibition of tumor necrosis factor production by the neuropeptide urocortin.

Urocortin (UCN) is a neuropeptide homologous with corticotropin-releasing factor (CRF), which has anti-inflammatory activities not all mediated by corticosteroids. In mice, UCN (1 microg/mouse sc) significantly reduced lipopolysaccharide (LPS)-induced serum tumor necrosis factor (TNF) and interleukin (IL)-1beta levels in vivo but did not affect serum IL-6. These effects were paralleled by a rise in corticosterone (CS) levels. Blockade of the CS increase by cyanoketone did not prevent TNF inhibition by UCN, suggesting the neuropeptide has anti-inflammatory mechanisms independent of the hypothalamus-pituitary-adrenal axis. In fact UCN had a direct inhibitory effect on LPS-induced TNF in rat Kupffer cells at concentrations between 10(-10) and 10(-16) M, and this effect was related to increased cAMP levels. However, the in vivo inhibition of LPS-induced IL-1beta by UCN was reversed by cyanoketone, indicating that the increase of endogenous glucocorticoids might be more important in IL-1beta inhibition than in TNF inhibition by UCN.

Peripheral effects of centrally administered interleukin-1beta in mice in relation to its clearance from the brain into the blood and tissue distribution.

Administration of interleukin IL-1 induces acute-phase response and inhibition of gastric secretion more efficiently when administered intracerebroventricularly (i.c.v.) than when the same dose of IL-1 is administered systemically. In this study we describe the pharmacokinetics of IL-1beta, administered centrally or systemically, in the serum or in peripheral tissues. IL-1beta administered i.c.v. resulted in higher peak IL-1beta concentrations, and lasted longer, than intravenous (i.v.) or intraperitoneal (i.p.) administration. Higher IL-1beta levels in the liver and heart were observed after i. c.v. administration (compared to the i.p. or i.v. route). Our data suggest that centrally injected IL-1 induces higher circulating and hepatic IL-1 levels and contributes to the fact that the i.c.v. route of administration is particularly effective in inducing a liver acute-phase response.

Granulocyte colony-stimulating factor and antibiotics in the prophylaxis of a murine model of polymicrobial peritonitis and sepsis.

Infections that occur after intraabdominal surgery still cause considerable morbidity and mortality despite the administration of prophylactic antibiotics. Increasing the number of neutrophils may also be a prophylactic approach, and granulocyte colony-stimulating factor (G-CSF) has been found to be beneficial in different animal models of peritonitis and sepsis. It is the combination of G-CSF and antibiotics, however, that is clinically relevant. Treatment of mice with G-CSF that was started before cecal ligation and puncture and continued afterward with antibiotics improved survival, decreased splenic bacterial colony-forming units and serum tumor necrosis factor, and increased serum interleukin-10, compared with treatment with antibiotics alone or with saline. Compared with saline, antibiotics alone increased tumor necrosis factor and did not affect interleukin-10. Thus, G-CSF confers onto antibiotics beneficial antiinfectious and antiinflammatory properties. A prophylactic regimen combining G-CSF and antibiotics may help prevent severe infectious complications following intraabdominal surgery.

Leptin causes body weight loss in the absence of in vivo activities typical of cytokines of the IL-6 family.

To investigate if leptin shares in vivo activities with interleukin (IL)-6 family cytokines, it was tested in normal mice for the ability, after a single injection, to induce the acute-phase protein serum amyloid A, to potentiate the induction by IL-1 of serum corticosterone and IL-6, and to inhibit the induction by lipopolysaccharide of serum tumor necrosis factor and, after seven daily injections, to cause body weight loss and to change peripheral blood cell counts. At a 0.5 mg/kg dose, leptin caused body weight loss but did not show any of the other activities above. At a dose of 5 mg/kg, which also caused body weight loss, leptin potentiated the induction by IL-1 of serum corticosterone and IL-6 but did not show any other activity. In addition to causing body weight loss, leptin shows only some of the in vivo activities typical of IL-6 family cytokines and only if used at a dose that exceeds the one sufficient to affect body weight. In vivo, leptin seems to chiefly control body weight and not inflammatory or hematopoietic processes.

Proteins of rat serum V: adjuvant arthritis and its modulation by nonsteroidal anti-inflammatory drugs.

The effect of adjuvant arthritis (AA) on the pattern of rat serum proteins includes the upregulation of haptoglobin, orosomucoid, alpha2-macroglobulin, serine protease inhibitor-3, thiostatin, alpha1-antitrypsin, C-reactive protein, and the downregulation of kallikrein-binding protein, alpha1-inhibitor III, apolipoprotein A-I, alpha2-HS-glycoprotein, albumin, apolipoprotein A-IV, transthyretin and transferrin. Minor changes (+/- 20%) are observed for Gc-globulin, ceruloplasmin, and alpha1-macroglobulin. AA thus grossly resembles the acute inflammatory response elicited by the injection of turpentine, although the changes in the levels of negative acute-phase proteins (APP) are smaller in acute inflammation. Indomethacine and ibuprofen inhibit the effects of arthritis on the synthesis of rat serum proteins in different ways: The former is, on average, three times as effective as the latter. Each drug interferes differently with different proteins. In animals without AA, both nonsteroidal anti-inflammatory drugs (NSAID) mimic the inflammatory pattern to a certain extent, with more effect on the negative than on the positive APPs. Overall, the shifts in serum protein levels parallel changes in inflammatory parameters such as joint swelling and serum interleukin-6 (IL-6) activity. Protein quantitation after two-dimensional electrophoresis (2-DE) reveals some effects of the drugs per se which escape detection by other routine tests.

Erythropoietin crosses the blood-brain barrier to protect against experimental brain injury.

Erythropoietin (EPO), recognized for its central role in erythropoiesis, also mediates neuroprotection when the recombinant form (r-Hu-EPO) is directly injected into ischemic rodent brain. We observed abundant expression of the EPO receptor at brain capillaries, which could provide a route for circulating EPO to enter the brain. In confirmation of this hypothesis, systemic administration of r-Hu-EPO before or up to 6 h after focal brain ischemia reduced injury by approximately 50-75%. R-Hu-EPO also ameliorates the extent of concussive brain injury, the immune damage in experimental autoimmune encephalomyelitis, and the toxicity of kainate. Given r-Hu-EPO's excellent safety profile, clinical trials evaluating systemically administered r-Hu-EPO as a general neuroprotective treatment are warranted.

Within-patient variability of hormone and cytokine concentrations in heart failure.

Several studies on disease and treatment effects on neurohormones have been conducted with small numbers of patients, using one blood sample as representative of their states. The aim of this study was to assess the within-patient variability of plasma concentrations of several hormones and cytokines of recent interest, in patients with moderate heart failure and controlled stable background therapy over 3 weeks. Blood for neurohormone and cytokine assays was sampled in duplicate from 18 patients with moderate heart failure. After an initial visit, the patients were kept on stable therapy until the second blood sampling 21 +/- 3 days later. The plasma concentrations of several neurohormones (endothelin, renin, angiotensin II, aldosterone, norepinephrine) and cytokines (interleukin-6 (IL-6), interleukin-13 (IL-13), ciliary neurotrophic factor (CNTF), leukemia inhibitory factor (LIF) and soluble receptor type I of tumour necrosis factor-alpha, (sTNF-RI) were measured with immunochemical methods. Some cytokines (IL-13, CNTF and LIF) were not detected. Despite clinically satisfactory ACE inhibition, circulating angiotensin II and aldosterone levels were still elevated in some patients, suggesting aldosterone escape. The between-visit agreement of plasma concentrations measured in duplicate was less than 35% for all circulating factors, except renin which showed a higher variability throughout the 3-week study period.