Cytotoxicity of root canal irrigating solutions and photodynamic therapy using curcumin photosensitizer.

BACKGROUND: Photodynamic therapy (PDT) has shown satisfactory antibacterial effects. However, little information regarding the cytotoxicity potential of PDT using curcumin as a photosensitizer (PS) on fibroblasts are found. The aim of this in vitro study was to evaluate the cytotoxicity of root canal irrigating solutions and photodynamic therapy with curcumin PS on the L-929 cell line. METHODS: Healthy mouse skin fibroblast cells were distributed into the following 7 experimental groups: G1 - culture medium DMEM (control group); G2 - 0.9% sodium chloride; G3 - 2.5% sodium hypochlorite (NaOCl); G4 - 5% NaOCl; G5 - PDT with curcumin PS at 500 mg/L + blue LED; G6 - PDT with curcumin PS at 750 mg/L + blue LED; and G7 - PDT with curcumin PS at 1000 mg/L + blue LED. All experimental groups which underwent PDT action were submitted to blue LED for 4 min, with a wavelength of 480 nm and energy fluency of 75 J/cm². The cultures were maintained under standard cell culture conditions (37°C, 100% humidity, 5% CO2). Cell viability analysis was performed using the colorimetric method to evaluate the periods of 6, 24, and 48 h. Data were subjected to the Kruskal-Wallis test, followed by the Dunn test to compare groups and Friedman test to compare periods (α = 0.05). RESULTS: When comparing the periods, no significant differences were observed for any of the experimental groups analyzed (p > 0.05), except for the NaOCl2.5 group that exhibited higher cell viability at 6 h compared to the period of 48 h (p = 0.0489). In the comparisons of the experimental groups, there were no statistically significant differences between the control group compared to all disinfection protocols, regardless of the period evaluated (p > 0.05), except for the PDT + C1000 group that showed lower cell viability (P < 0.05). CONCLUSIONS: PDT with curcumin at 1000 mg/L was cytotoxic on L-929 fibroblast cell culture. However, laser-activated curcumin at a concentration of 500 mg/L presented no influence on L-929 fibroblast cell viability in in vitro conditions.

Arte de cuidar milenar: crenças e saberes de idosos sobre a fitoterapia / Arte milenario de cuidar: creencias y saberes de los adultos mayores sobre la fitoterapia / Millennial art of caring: beliefs and knowledge of the elderly about phytotherapy

Existe uma vasta gama de terapias conhecidas no mundo e uma das principais, que faz parte de diversas culturas no mundo percorrendo a história da humanidade, é a utilização das plantas e ervas medicinais com finalidade terapêutica. No Brasil, deduz-se que as práticas de fitoterapia estejam presentes desde antes da colonização, quando os indígenas faziam o uso curativo de acordo com seus conhecimentos e suas experiências [Fragmento de texto] (AU)

l-arginine modulates inflammation and muscle regulatory genes after a single session of resistance exercise in rats.

We investigated the skeletal muscle adaptation to l-arginine supplementation prior to a single session of resistance exercise (RE) during the early phase of muscle repair. Wistar rats were randomly assigned into non-exercised (Control), RE plus vehicle (RE); RE plus l-arginine (RE+L-arg) and RE plus aminoguanidine (RE+AG) groups. Animals received four doses of either vehicle (0.9% NaCl), l-arg (1 g/b.w.), or AG (iNOS inhibitor) (50 mg/b.w.). The animals performed a single RE session until the concentric failure (ladder climbing; 80% overload) and the skeletal muscles were harvested at 0, 8, 24, and 48 hours post-RE. The RE resulted in increased neutrophil infiltrate (24 hours post-RE) (3621 vs 11852; P<.0001) associated with enhanced TNF-α (819.49 vs 357.02; P<.005) and IL-6 (3.84 vs 1.08; P<.0001). Prior, l-arginine supplementation attenuates neutrophil infiltration (5622; P<.0001), and also TNF-α (506.01; P<.05) and IL-6 (2.51, P<.05) levels. AG pretreatment mediated an inhibition of iNOS levels similar to levels found in RE group. RE animals displayed increased of atrogin-1 (1.9 fold) and MuRF-1 (3.2 fold) mRNA levels, reversed by l-arg supplementation [atrogin-1 (0.6 fold; P<.001); MuRF-1 (0.8-fold; P<.001)] at 24 hours post-RE. MyoD up-regulated levels were restricted to l-arg treated animals at 24 hours (2.8 vs 1.5 fold; P<.005) and 48 hours post-RE (2.4 vs 1.1 fold; P<.001). AG pretreatment reversed these processes at 24 hours [atrogin-1 (2.1 fold; P<.0001); MuRF-1 (2.5 fold; P<.0001); MyoD (1.4 fold)]. l-arginine supplementation seems to attenuate the resolution of RE-induced muscle inflammation and up-regulates MyoD expression during the early phase of muscle repair.

The influence of implant location and position characteristics on peri-implant pathology..

Peri-implant pathology is a multifactorial disease, incorporating biological and biomechanical components in its pathogenesis; however; few studies address the possible risk factors. This study investigated the effect of implant location and position characteristics on the occurrence of Peri-implant pathology. A total of 1350 patients with dental implants were included 270 patients with peri-implant pathology and 1080 healthy controls. Results demonstrated that in the absence of bacterial plaque and smoking, the variable proximity of the implant to other implants or teeth revealed a significant difference between groups with a protective effect, but not in the presence of bacterial plaque and smoking.

Hypertension favors the inflammatory process in rats with experimentally induced periodontitis.

BACKGROUND AND OBJECTIVE: Cardiovascular diseases are significantly correlated with chronic periodontitis. The aim of this study was to evaluate bone-loss level, neutrophil migration, CXCL2/CINC-2α, CXCL5/LIX, CCL20/MIP-3α and tumor necrosis factor-α (TNF-α) production, inducible nitric oxide synthase (iNOS) expression and C-reactive protein (CRP) release in spontaneously hypertensive rats (SHRs) and normotensive (WTK) rats after experimental induction of periodontal disease. MATERIAL AND METHODS: Periodontitis was induced by placement of silk yarn ligatures around the first molar counterparts. The levels of CRP, CCL20/MIP-3α and CXCL5/LIX were evaluated in the peripheral blood, and bone-loss level, neutrophil recruitment, the production of myeloperoxidase, CXCL2, CXCL5, CCL20 and TNF-α, and the expression of iNOS were evaluated in the gingival tissue. Histological sections were taken to evaluate and measure bone resorption and neutrophil recruitment in the furcation region. RESULTS: Rats with periodontitis had alveolar bone resorption. SHRs with periodontitis showed marked bone loss and increased neutrophil infiltration in comparison with WTK rats. SHRs with periodontitis showed increased levels of TNF-α and CXCL2, and a slight tendency for increased levels of CXCL5, in the gingival tissue but no increase in the level of CCL20. In SHRs, even without periodontitis, the levels of TNF-α, CXCL2, CXCL5 and CCL20 showed a slight tendency to increase. In the WTK rats, TNF-α, CXCL2 and CXCL5 levels were increased with periodontitis, but the level of CCL20 was not. iNOS was expressed in the gingival tissue of WTK rats and SHRs with periodontitis; however, SHRs appeared to express a higher level of iNOS than did WKT rats. The CRP level was elevated in both types of rats with periodontitis; however, the CRP level was higher in SHRs with periodontitis than in WTK rats with periodontitis. CONCLUSION: In SHRs, the hypertensive condition per se seems to favor the inflammatory processes that become potentiated with periodontitis, when compared with WKT rats.

Characterization of a local renin-angiotensin system in rat gingival tissue.

BACKGROUND: The systemic renin-angiotensin system (RAS) promotes the plasmatic production of angiotensin (Ang) II, which acts through interaction with specific receptors. There is growing evidence that local systems in various tissues and organs are capable of generating angiotensins independently of circulating RAS. The aims of this study were to investigate the expression and localization of RAS components in rat gingival tissue and evaluate the in vitro production of Ang II and other peptides catalyzed by rat gingival tissue homogenates incubated with different Ang II precursors. METHODS: Reverse transcription-polymerase chain reaction assessed mRNA expression. Immunohistochemical analysis aimed to detect and localize renin. A standardized fluorimetric method with tripeptide hippuryl-histidyl-leucine was used to measure tissue angiotensin-converting enzyme (ACE) activity, whereas high performance liquid chromatography showed products formed after the incubation of tissue homogenates with Ang I or tetradecapeptide renin substrate (TDP). RESULTS: mRNA for renin, angiotensinogen, ACE, and Ang II receptors (AT(1a), AT(1b), and AT(2)) was detected in gingival tissue; cultured gingival fibroblasts expressed renin, angiotensinogen, and AT(1a) receptor. Renin was present in the vascular endothelium and was intensely expressed in the epithelial basal layer of periodontally affected gingival tissue. ACE activity was detected (4.95 +/- 0.89 nmol histidyl-leucine/g/minute). When Ang I was used as substrate, Ang 1-9 (0.576 +/- 0.128 nmol/mg/minute), Ang II (0.066 +/- 0.008 nmol/mg/minute), and Ang 1-7 (0.111 +/- 0.017 nmol/mg/minute) were formed, whereas these same peptides (0.139 +/- 0.031, 0.206 +/- 0.046, and 0.039 +/- 0.007 nmol/mg/minute, respectively) and Ang I (0.973 +/- 0.139 nmol/mg/minute) were formed when TDP was the substrate. CONCLUSION: Local RAS exists in rat gingival tissue and is capable of generating Ang II and other vasoactive peptides in vitro.

Neutrophil migration induced by IL-1beta depends upon LTB4 released by macrophages and upon TNF-alpha and IL-1beta released by mast cells.

In the present study, we investigate whether mast cells and macrophages are involved in the control of IL-1beta-induced neutrophil migration, as well as the participation of chemotactic mediators. IL-1beta induced a dose-dependent neutrophil migration to the peritoneal cavity of rats which depends on LTB(4), PAF and cytokines, since the animal treatment with inhibitors of these mediators (MK 886, PCA 4248 and dexamethasone respectively) inhibited IL-1beta-induced neutrophil migration. The neutrophil migration induced by IL-1beta is dependent on mast cells and macrophages, since depletion of mast cells reduced the process whereas the increase of macrophage population enhanced the migration. Moreover, mast cells or macrophages stimulated with IL-1beta released a neutrophil chemotactic factor, which mimicked the neutrophil migration induced by IL-1beta. The chemotactic activity of the supernatant of IL-1beta-stimulated macrophages is due to the presence of LTB(4), since MK 886 inhibited its release. Moreover, the chemotactic activity of IL-1beta-stimulated mast cells supernatant is due to the presence of IL-1beta and TNF-alpha, since antibodies against these cytokines inhibited its activity. Furthermore, significant amounts of these cytokines were detected in the supernatant. In conclusion, our results suggest that neutrophil migration induced by IL-1beta depends upon LTB(4) released by macrophages and upon IL-1beta and TNFalpha released by mast cells.

Molecular mechanisms mediating gastrin-releasing peptide receptor modulation of memory consolidation in the hippocampus.

Although the gastrin-releasing peptide-preferring bombesin receptor (GRPR) has been implicated in memory formation, the underlying molecular events are poorly understood. In the present study, we examined interactions between the GRPR and cellular signaling pathways in influencing memory consolidation in the hippocampus. Male Wistar rats received bilateral infusions of bombesin (BB) into the dorsal hippocampus immediately after inhibitory avoidance (IA) training. Intermediate doses of BB enhanced, whereas a higher dose impaired, 24-h IA memory retention. The BB-induced memory enhancement was prevented by pretraining infusions of a GRPR antagonist or inhibitors of protein kinase C (PKC), mitogen-activated protein kinase (MAPK) kinase and protein kinase A (PKA), but not by a neuromedin B receptor (NMBR) antagonist. We next further investigated the interactions between the GRPR and the PKA pathway. BB-induced enhancement of consolidation was potentiated by coinfusion of activators of the dopamine D1/D5 receptor (D1R)/cAMP/PKA pathway and prevented by a PKA inhibitor. We conclude that memory modulation by hippocampal GRPRs is mediated by the PKC, MAPK, and PKA pathways. Furthermore, pretraining infusion of BB prevented beta-amyloid peptide (25-35)-induced memory impairment, supporting the view that the GRPR is a target for the development of cognitive enhancers for dementia.

Interleukin 4 induces the expression of inducible nitric oxide synthase in eosinophils.

We investigated the effects of the Th2-like cytokines IL-4 and IL-13 and of IL-10 on the induction of iNOS and NO production in rat eosinophils. Addition of mIL-4 to the eosinophil culture increased iNOS activity and nitrite production but did not improve the stimulatory effect of IFN-gamma and LPS. In contrast to eosinophils, addition of mIL-4 to macrophage cultures inhibited the iNOS expression and nitrite production induced by IFN-gamma plus LPS. Addition of mIL-13 to the eosinophil cultures did not significantly change iNOS activity and nitrite production in cells stimulated or not with IFN-gamma plus LPS. On the other hand, IL-13 inhibited iNOS activity in IFN-gamma plus LPS-stimulated macrophages. In the presence of IL-10, iNOS activity in non-stimulated eosinophil or macrophage cultures was not significantly altered, but the enzyme expression was inhibited in IFN-gamma plus LPS-stimulated eosinophils or macrophages. The production of nitrite by eosinophils stimulated by IFN-gamma plus LPS was inhibited by the presence of IL-10 in the medium. In conclusion, eosinophils might exhibit differential modulation of the L-arginine/iNOS pathway depending on the profile of Th2 cytokines produced during allergic diseases. IL-4 appears to be an important Th2 cytokine involved in the induction of the L-arginine/iNOS pathway in eosinophils.

Macrophages and mast cells control the neutrophil migration induced by dentin proteins.

Dentin sialoprotein (DSP) and dentin phosphoprotein (DPP), the major dentin proteins, have been shown to induce neutrophil migration through release of IL-1beta, TNF-alpha, MIP-2, and KC. However, the sources of these mediators were not determined. Here, the roles of macrophages and mast cells (MC) in dentin-induced neutrophil accumulation were investigated. Peritoneal MC depletion or the enhancement of macrophage population increased DSP- and DPP-induced neutrophil extravasation. Moreover, supernatants from DSP- and DPP-stimulated macrophages caused neutrophil migration. The release of neutrophil chemotactic factor by macrophages was inhibited by dexamethasone or the supernatant of DSP-treated MC. Consistently, dexamethasone and the MC supernatant inhibited the production of IL-1beta, TNF-alpha, and MIP-2 by macrophages. This inhibitory activity of the DSP-stimulated MC was neutralized by anti-IL-4 and anti-IL-10 antibodies. These results indicate that dentin induces the release of the neutrophil chemotactic substance(s) by macrophages, which are down-modulated by MC-derived IL-4 and IL-10.

The role of chemokines and chemokine receptors in eosinophil activation during inflammatory allergic reactions

Chemokines are important chemotactic cytokines that play a fundamental role in the trafficking of leukocytes to sites of inflammation. They are also potent cell-activating factors, inducing cytokine and histamine release and free radical production, a fact that makes them particularly important in the pathogenesis of allergic inflammation. The action of chemokines is regulated at the level of agonist production and processing as well as at the level of receptor expression and coupling. Therefore, an analysis of the ligands must necessarily consider receptors. Eosinophils are target cells involved in the allergic inflammatory response since they are able to release a wide variety of mediators including CC and CXC chemokines and express their receptors. These mediators could damage the airway epithelial cells and might be important to stimulate other cells inducing an amplification of the allergic response. This review focuses on recently emerging data pertaining to the importance of chemokines and chemokine receptors in promoting eosinophil activation and migration during the allergic inflammatory process. The analysis of the function of eosinophils and their chemokine receptors during allergic inflammation might be a good approach to understanding the determinants of asthma severity and to developing novel therapies.

The role of chemokines and chemokine receptors in eosinophil activation during inflammatory allergic reactions.

Chemokines are important chemotactic cytokines that play a fundamental role in the trafficking of leukocytes to sites of inflammation. They are also potent cell-activating factors, inducing cytokine and histamine release and free radical production, a fact that makes them particularly important in the pathogenesis of allergic inflammation. The action of chemokines is regulated at the level of agonist production and processing as well as at the level of receptor expression and coupling. Therefore, an analysis of the ligands must necessarily consider receptors. Eosinophils are target cells involved in the allergic inflammatory response since they are able to release a wide variety of mediators including CC and CXC chemokines and express their receptors. These mediators could damage the airway epithelial cells and might be important to stimulate other cells inducing an amplification of the allergic response. This review focuses on recently emerging data pertaining to the importance of chemokines and chemokine receptors in promoting eosinophil activation and migration during the allergic inflammatory process. The analysis of the function of eosinophils and their chemokine receptors during allergic inflammation might be a good approach to understanding the determinants of asthma severity and to developing novel therapies.

Stem cell factor: a hemopoietic cytokine with important targets in asthma.

We review evidence that Stem Cell Factor (SCF) plays an important role in the pathophysiology of asthma. SCF is produced by a wide variety of cells present in asthmatic lung, including mast cells and eosinophils. Its receptor, c-kit, is broadly expressed on mature mast cells and eosinophils. SCF promotes recruitment of mast cell progenitors into tissues, as well as their local maturation and activation. It also promotes eosinophil survival, maturation and functional activation. SCF enhances IgE-dependent release of mediators from mast cells, including histamine, leukotrienes, cytokines (TNF-alpha, IL-5, GM-CSF) and chemokines (RANTES/CCL5, MCP-1/CCL2, TARC/CCL17 e MDC/CCL22); it is required for IL-4 production in mast cells. SCF, acting in concert with IgE, also upregulates the expression and function of CC chemokine receptors in mast cells. Structural and resident airway cells express increased levels of SCF in the bronchus of asthmatic patients. In a murine model of asthma, allergen exposure increased production of SCF by epithelial cells and alveolar macrophages, which was transient and paralleled by histamine release. SCF induced long-lived airway hyperreactivity, which was prevented by local neutralization of SCF, as well as by inhibitors of the production or activity of cysteinyl-leukotrienes. Together, these observations suggest that SCF has an important role in asthma.

Sephadex induces eosinophil migration to the rat and mouse peritoneal cavity: involvement of mast cells, LTB4, TNF-alpha, IL-8 and PAF.

OBJECTIVE AND DESIGN: In this study we investigated the chemotactic mediators involved in the Sephadex-induced eosinophil migration into the peritoneal cavities of rats and mice, and which resident peritoneal cells release these mediators. MATERIALS AND METHODS: Sephadex suspension was injected into the peritoneal cavities of rats or mice which were pretreated, or not, with specific drugs that inhibit synthesis or production of the inflammatory mediators and eosinophil chemotactic activities were observed. To investigate the role of resident peritoneal cells as a source of these chemotactic factors, the macrophage population was enhanced or the mast cell population was depleted. The resident cells were also stimulated, in vitro, with Sephadex and the chemotactic activity of the supernatants was determined. RESULTS: Sephadex induced dose and time dependent eosinophil migration in rats and mouse, which were inhibited by dexamethasone and MK 886. BN 52021 only affected the eosinophil migration into the mouse peritoneal cavity. An increase in the macrophage population did not alter the eosinophil migration induced by Sephadex in rat or mouse. However, mast cell population depletion reduced eosinophil migration in rats, but did not alter the migration in mice. Sephadex-stimulated rat mast cells released an eosinophil chemotactic factor whose release was inhibited by dexamethasone and MK 886. Anti-TNF-alpha and anti-IL-8 Abs inhibited the chemotactic activity of the mast cell supernatant. CONCLUSION: Sephadex-induced eosinophil migration into the rat peritoneal cavity is dependent on mast cells, which release LTB4, TNF-alpha and CINC-1. Conversely, Sephadex-induced eosinophil migration into the mouse peritoneal cavity is mediated by PAF and LTB4, which are not released from resident macrophages or mast cells.

Stem cell factor and igE-stimulated murine mast cells produce chemokines (CCL2, CCL17, CCL22) and express chemokine receptors.

OBJECTIVE AND DESIGN: In the present study we investigated the effect of SCF and/or IgE on histamine, TNF-alpha and chemokines released from bone marrow-derived mast cells (BMMC) as well as chemokine receptor expression. MATERIAL AND METHODS: BMMC were derived from femoral bone marrow of CBA/J mice. The purity of BMMC was >98% after 3 weeks. BMMC (2.5 x 10(6) cells/well) were incubated in the presence or absence of either SCF, IgE plus DNP or a combination of SCF and IgE for 6 and 18 h. Cell-free supernatants were recovered to measure CC chemokines, TNF-alpha and histamine release utilizing ELISA assays. CC chemokine family receptors were detected by RT-PCR analysis, and confirmed using functional chemotactic assays. RESULTS: Histamine levels were comparable between SCF and IgE stimulated cells, whereas TNF-alpha production was significantly greater after IgE compared to SCF stimulation. SCF and/or IgE-stimulated BMMC released CC chemokines, CCL22 (MDC), CCL17 (TARC) and CCL2 (MCP-1). Increased mRNA expression of CCR1, CCR2, CCR3, and CCR5 was detected in SCF and IgE-stimulated BMMCs. Functional chemotactic assays confirmed the expression data. CONCLUSION: SCF and IgE can up-regulate the expression of chemokines and chemokine receptors on mast cells. Thus, SCF may play a significant role in their activation and inflammation during allergic responses.

SCF-induced airway hyperreactivity is dependent on leukotriene production.

Stem cell factor (SCF) is directly involved in the induction of airway hyperreactivity during allergen-induced pulmonary responses in mouse models. In these studies, we examined the specific mediators and mechanisms by which SCF can directly induce airway hyperreactivity via mast cell activation. Initial in vitro studies with bone marrow-derived mast cells indicated that SCF was able to induce the production of bronchospastic leukotrienes, LTC(4) and LTE(4). Subsequently, when SCF was instilled in the airways of naive mice, we were able to observe a similar induction of LTC(4) and LTE(4) in the bronchoalveolar lavage (BAL) fluid and lungs of treated mice. These in vivo studies clearly suggested that the previously observed SCF-induced airway hyperreactivity may be related to the leukotriene production after SCF stimulation. To further investigate whether the released leukotrienes were the mediators of the SCF-induced airway hyperreactivity, an inhibitor of 5-lipoxygenase (5-LO) binding to the 5-LO activating protein (FLAP) was utilized. The FLAP inhibitor MK-886, given to the animals before intratracheal SCF administration, significantly inhibited the release of LTC(4) and LTE(4) into the BAL fluid. More importantly, use of the FLAP inhibitor nearly abrogated the SCF-induced airway hyperreactivity. In addition, blocking the LTD(4)/E(4), but not LTB(4), receptor attenuated the SCF-induced airway hyperreactivity. In addition, the FLAP inhibitor reduced other mast-derived mediators, including histamine and tumor necrosis factor. Altogether, these studies indicate that SCF-induced airway hyperreactivity is dependent upon leukotriene-mediated pathways.

Novel protective effects of stem cell factor in a murine model of acute septic peritonitis. Dependence on MCP-1.

Mast cells participate in the host response during sepsis and have been shown to have a protective effect in a murine model of acute septic peritonitis and multi-organ failure initiated by cecal ligation and puncture (CLP). Stem cell factor (SCF) is a hematopoietic cytokine important in mast cell proliferation and activation. In the present study, we examined the protective effects of a single intraperitoneal injection of SCF given 2 hours before CLP surgery in mice. Four days after the CLP surgery, SCF pretreatment significantly improved mouse survival from 29 to 56% and mast cells were absolutely required for this effect. Immunoneutralization studies revealed that the SCF-stimulated release of monocyte chemoattractant protein-1 (MCP-1) into the septic peritoneal cavity contributed to the protective effect of SCF in this model. One potential cellular source of MCP-1 was the SCF-activated mast cell. In addition, SCF pretreatment significantly augmented circulating levels of SCF and the immunomodulatory cytokine interleukin-10 in septic mice, in part because the SCF pretreatment seemed to promote the release of both mediators from the liver. Additional hepatic effects of SCF treatment included an accelerated expression of hepatic levels of signal transducer and activator of transcription-3 (STAT-3) in CLP mice pretreated with SCF. Taken together, the findings from the present study demonstrate that the intraperitoneal delivery of SCF has a major protective effect in a murine model of CLP.

Favourable outcome in 135 children with juvenile systemic sclerosis: results of a multi-national survey.

OBJECTIVE: To increase the current knowledge of the outcome of juvenile systemic sclerosis (jSSc), which is currently limited. METHODS: In order to investigate the patient outcome and prognostic factors, starting October 1994, we distributed questionnaires to 324 paediatric rheumatology centres. RESULTS: Until 15 May 1998 responses from 46 centres were received, 34 of which returned completed questionnaires on a total of 135 patients. One hundred and twenty-two of the 135 patients were Caucasian, 100 were female. The mean age at disease onset was 8.8 yr (S.D. +/- 3.3 yr). The mean disease duration at the last follow-up was 5 yr(S.D. +/- 3.3 yr). At the last follow-up the disease was still active and required medication in 82 patients, 36 had inactive disease on medication, and 16 were in remission. Ninety per cent of the living patients were fully active in daily life at the last follow-up. Eight of the 135 patients had died. These patients had a median age at onset of the disease of 10.5 yr (range 6.7-15.8 yr). The median disease duration until death was 2 yr (range 1-8 yr). The causes of death were heart failure (five), renal failure (one), sepsis (one) and in one case the cause was not defined. The 1 yr survival rate was 99%, the 2 yr was 97% and the 4 yr was 95%. CONCLUSIONS: At a mean follow-up of 5 yr, the current results show a favourable outcome in most patients with childhood onset jSSc and a significantly better survival than in the adult SSc patients.

Microbicidal activity of eosinophils is associated with activation of the arginine-NO pathway.

In order to investigate the ability of rat peritoneal eosinophils to produce nitric oxide (NO) induced by cytokines in vitro, these cells were activated with several cytokines (IL-5, IL-8, Rantes, TNF-alpha, IFN-gamma) in association or not with LPS. Under these conditions, we were able to detect nitrite in the incubation medium when the eosinophils were stimulated with IFN-gamma or IL-8 in the presence of LPS. LPS alone also induced nitrite production. Significant levels of nitrite in the medium were already present after 12 h of stimulation and increased steadily within the next 48 h. Regarding NO synthase, its highest activity was achieved at 12 h after IFN-gamma/LPS stimulation. After this peak, the enzymatic activity reduced gradually to control levels 48 h after the stimulation. The simultaneous addition of the NO synthase inhibitor L-NIO (100 microM) to the eosinophil suspension blocked nitrite production and NO synthase activity. On the other hand, neither IL-5, Rantes nor TNF-alpha were able to induce the release of nitrite in the presence or absence of LPS. To evaluate the microbicidal effect of these cells against the Leishmania parasite, eosinophils were infected with Leishmania major. It was observed that these cells were able to produce nitrite and to kill the parasite after activation with LPS/IFN-gamma. Moreover, L-NIO blocked this leishmanicidal activity and the nitrite production. Our results suggest that activated eosinophils release NO which is involved in their microbicidal activity against Leishmania major.