Talc pleurodesis: Evidence of systemic Inflammatory response to small size talc particles

The mechanisms of the systemic response associated with talc-induced pleurodesis are poorly understood. The aim of this study was to assess the acute inflammatory response and migration of talc of small. size particles injected in the pleural space. Rabbits were injected intrapleurally with talc solution containing small. or mixed particles and blood and pleural fluid samples were collected after 6, 24 or 48 h and assayed for leukocytes, neutrophils, lactate dehydrogenase, IL-8, VEGF, and TGF-beta. The lungs, spleen, liver and kidneys were assessed to study deposit of talc particles. Both types of talc produced an acute serum inflammatory response, more pronounced in the small particles group. Pleural fluid IL-8 and VEGF levels were higher in the small particle talc group. Correlation between pleural VEFG and TGF-beta levels was observed for both groups. Although talc particles were demonstrated in the organs of both groups, they were more pronounced in the small talc group. In conclusion, intrapleural injection of talc of small size particles produced a more pronounced acute systemic response and a greater deposition in organs than talc of mixed particles. (C) 2008 Elsevier Ltd. All rights reserved.

Elevated striatal and decreased dorsolateral prefrontal cortical activity in response to emotional stimuli in euthymic bipolar disorder: no associations with psychotropic medication load

To examine abnormal patterns of frontal cortical-subcortical activity in response to emotional stimuli in euthymic individuals with bipolar disorder type I in order to identify trait-like, pathophysiologic mechanisms of the disorder. We examined potential confounding effects of total psychotropic medication load and illness variables upon neural abnormalities. We analyzed neural activity in 19 euthymic bipolar and 24 healthy individuals to mild and intense happy, fearful and neutral faces. Relative to healthy individuals, bipolar subjects had significantly increased left striatal activity in response to mild happy faces (p < 0.05, corrected), decreased right dorsolateral prefrontal cortical (DLPFC) activity in response to neutral, mild and intense happy faces, and decreased left DLPFC activity in response to neutral, mild and intense fearful faces (p < 0.05, corrected). Bipolar and healthy individuals did not differ in amygdala activity in response to either emotion. In bipolar individuals, there was no significant association between medication load and abnormal activity in these regions, but a negative relationship between age of illness onset and amygdala activity in response to mild fearful faces (p = 0.007). Relative to those without comorbidities, bipolar individuals with comorbidities showed a trend increase in left striatal activity in response to mild happy faces. Abnormally increased striatal activity in response to potentially rewarding stimuli and decreased DLPFC activity in response to other emotionally salient stimuli may underlie mood instabilities in euthymic bipolar individuals, and are more apparent in those with comorbid diagnoses. No relationship between medication load and abnormal neural activity in bipolar individuals suggests that our findings may reflect pathophysiologic mechanisms of the illness rather than medication confounds. Future studies should examine whether this pattern of abnormal neural activity could distinguish bipolar from unipolar depression.

CCR2 receptor is essential to activate microbicidal mechanisms to control Toxoplasma gondii infection in the central nervous system

Chemokines comprise a structurally related family of cytokines that regulate leukocyte trafficking. Because infection with Toxoplasma gondii can induce an important inflammatory reaction that, if left uncontrolled, can lead to death, we investigated the role of the chemokine receptor CCR2 in T gondii infection. We orally infected CCR2(-/-) mice with five ME-49 T gondii cysts and monitored morbidity, survival, and immune response thereafter. The CCR2(-/-) mice displayed higher susceptibility to infection as all mice died on day 28 after infection. Despite similar Th1 responses, a more evident anti-inflammatory response was induced in the peripheral organs of CCR2(-/-) mice compared with wild-type C57BL/6 mice. Additionally, CCR2-/- mice presented greater parasitism and a milder inflammatory reaction in their peripheral organs with lesser CD4(+) and MAC-1(+) and greater CD8(+) cell migration. The parasite load decreased in these organs in CCR2(-/-) mice but remained uncontrolled in the central nervous system. Additionally, we observed down-regulated inducible nitric oxide synthase expression in peripheral organs from CCR2(-/-) mice that was associated with a small nitric oxide production by spleen macrophages. In conclusion, in the absence of CCR2, another mechanism is activated to control tissue parasitism in peripheral organs. Nevertheless, CCR2 is essential for the activation of microbicidal mediators that control T gondii replication in the central nervous system.

Host genetic background affects regulatory T-cell activity that influences the magnitude of cellular immune response against Mycobacterium tuberculosis

Using two mouse strains with different abilities to generate interferon (IFN)-gamma production after Mycobacterium tuberculosis infection, we tested the hypothesis that the frequency and activity of regulatory T (Treg) cells are influenced by genetic background. Our results demonstrated that the suppressive activity of spleen Treg cells from infected or uninfected BALB/c mice was enhanced, inhibiting IFN-gamma and interleukin (IL)-2 production. Infected C57BL/6 mice exhibited a decrease in the frequency of lung Treg cells and an increased ratio CD4(+):CD4(+)Foxp3(+) cells compared with infected BALB/c mice and uninfected C57BL/6 mice. Moreover, infected C57BL/6 mice also had a decrease in the immunosuppressive capacity of spleen Treg cells, higher lung IFN-gamma and IL-17 production, and restricted the infection better than BALB/c mice. Adoptive transfer of BALB/c Treg cells into BALB/c mice induced an increase in bacterial colony-forming unit (CFU) counts. Furthermore, BALB/c mice treated with anti-CD25 antibody exhibited lung CFU counts significantly lower than mice treated with irrelevant antibody. Our results show that in BALB/c mice, the Treg cells have a stronger influence than that in C57BL/6 mice. These data suggest that BALB/c and C57BL/6 mice may use some different mechanisms to control M. tuberculosis infection. Therefore, the role of Treg cells should be explored during the development of immune modulators, both from the perspective of the pathogen and the host. Immunology and Cell Biology (2011) 89, 526-534; doi:10.1038/icb.2010.116; published online 19 October 2010

Pore Formation Triggered by Legionella spp. Is an Nlrc4 Inflammasome-Dependent Host Cell Response That Precedes Pyroptosis

Legionella pneumophila, the etiological agent of Legionnaires disease, is known to trigger pore formation in bone marrow-derived macrophages (BMMs) by mechanisms dependent on the type IVB secretion system known as Dot/Icm. Here, we used several mutants of L. pneumophila in combination with knockout mice to assess the host and bacterial factors involved in pore formation in BMMs. We found that regardless of Dot/Icm activity, pore formation does not occur in BMMs deficient in caspase-1 and Nlrc4/Ipaf. Pore formation was temporally associated with interleukin-1 beta secretion and preceded host cell lysis and pyroptosis. Pore-forming ability was dependent on bacterial Dot/Icm but independent of several effector proteins, multiplication, and de novo protein synthesis. Flagellin, which is known to trigger the Nlrc4 inflammasome, was required for pore formation as flaA mutant bacteria failed to induce cell permeabilization. Accordingly, transfection of purified flagellin was sufficient to trigger pore formation independent of infection. By using 11 different Legionella species, we found robust pore formation in response to L. micdadei, L. bozemanii, L. gratiana, L. jordanis, and L. rubrilucens, and this trait correlated with flagellin expression by these species. Together, the results suggest that pore formation is neither L. pneumophila specific nor the result of membrane damage induced by Dot/Icm activity; instead, it is a highly coordinated host cell response dependent on host Nlrc4 and caspase-1 and on bacterial flagellin and type IV secretion system.

Mechanisms underlying the biphasic effect of vitamin K-1 (phylloquinone) on arterial blood pressure

Phylloquinone (vitamin K-1, VK1) is widely used therapeutically and intravenous administration of this quinone can induce hypotension. We aimed to investigate the mechanisms underlying the effects induced by VK1 on arterial blood pressure. With this purpose a catheter was inserted into the abdominal aorta of male Wistar rats for blood pressure and heart rate recording. Bolus intravenous injection of VK1 (0.5-20 mg kg(-1)) produced a transient increase in blood pressure followed by a fall. Both the pressor and depressor response induced by VK1 were dose-dependent. On the other hand, intravenous injection of VK1 did not alter heart rate. The nitric oxide synthase (NOS) inhibitor N-G-nitro-L-arginine methyl ester (L-NAME, 10 and 20 mg kg(-1)) reduced both the increase and decrease in blood pressure induced by VK1 (5 mgkg(-1)). On the other hand, indometacin (10 mg kg(-1)), a non-selective cyclooxygenase inhibitor, did not alter the increase in mean arterial pressure (MAP) induced by VK1. However, VK1-induced fall in MAP was significantly attenuated by indometacin. We concluded that VK1 induces a dose-dependent effect on blood pressure that consists of an acute increase followed by a more sustained decrease in MAP. The hypotension induced by VK1 involves the activation of the nitric oxide (NO) pathway and the release of vasodilator prostanoid(s).

Paraventricular nucleus mediates pressor response to noradrenaline injection into the dorsal periaqueductal gray area

The dorsal periaqueductal gray area (dPAG) is involved in cardiovascular modulation. In a previous study, we reported that noradrenaline (NA) microinjection into the dPAG of rats caused pressor response that was mediated by vasopressin release. Vasopressin is synthesized by magnocellular neurons in the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei. In the present study, we verified which nuclei mediated the cardiovascular response to NA as well as the existence of direct neural projection from the dPAG to hypothalamic nuclei. Then, we studied the effect of treating either PVN or SON with the nonselective synaptic blocker cobalt chloride (1 mM) on the cardiovascular response to NA (15 nmol) microinjection into dPAG. Attempting to identify neural projections from dPAG to hypothalamic nuclei, we microinjected the neuronal tracer biotinylated-dextran-amine (BDA) into the dPAG and searched varicosity-containing nerve terminals in the PVN and SON. Unilateral cobalt-induced inhibition of synapses in the SON did not affect the cardiovascular response to NA. However, unilateral inhibition of PVN significantly reduced the pressor response to NA. Moreover, cobalt-induced inhibition of synapses in both PVN blocked the pressor response caused by NA microinjected into the dPAG. Microinjection of BDA into the dPAG evidenced presence of varicosity-containing neuronal fibers in PVN but not in SON. The results from cobalt treatment indicated that synapses in PVN mediate the vasopressin-induced pressor response caused by NA microinjection into the dPAG. In addition, the neuroanatomical results from BDA microinjection into the dPAG pointed out the existence of direct neural projections from the dPAG site to the PVN. (C) 2009 Elsevier B.V. All rights reserved.

Lateral Parabrachial Afferent Areas and Serotonin Mechanisms Activated by Volume Expansion

Recent evidence has shown that the serotonergic mechanism of the lateral parabrachial nucleus (LPBN) participates in the regulation of renal and hormonal responses to isotonic blood volume expansion (BVE). We investigated the BVE-induced Fos activation along forebrain and hindbrain nuclei and particularly within the serotonergic clusters of the raphe system that directly project to the LPBN. We also examined whether there are changes in the concentration of serotonin (5HT) within the raphe nucleus in response to the same stimulus. With this purpose, we analyzed the cells doubly labeled for Fos and Fluorogold (FG) following BVE (NaCl 0.15 M, 2 ml/100 g b.w., 1 min) 7 days after FG injection into the LPBN. Compared with the control group, blood volume-expanded rats showed a significant greater number of Fos-FG double-labeled cells along the nucleus of the solitary tract, locus coeruleus, hypothalamic paraventricular nucleus, central extended amygdala complex, and dorsal raphe nucleus (DRN) cells. Our study also showed an increase in the number of serotonergic DRN neurons activated in response to isotonic BVE. We also observed decreased levels of 5HT and its metabolite 5-hydroxyindoleacetic acid (measured by high-pressure liquid chromatography) within the raphe nucleus 15 min after BVE. Given our previous evidence on the role of the serotonergic system in the LPBN after BVE, the present morphofunctional findings suggest the existence of a key pathway (DRN-LPBN) that may control BVE response through the modulation of 5HT release. (c) 2008 Wiley-Liss, Inc.

Glutamatergic and purinergic mechanisms on respiratory modulation in the caudal NTS of awake rats

In the present study we evaluated the role of ionotropic glutamate receptors and purinergic P2 receptors in the caudal commissural NTS (cNTS) on the modulation of the baseline respiratory frequency (fR), and on the tachypneic response to chemoreflex activation in awake rats. The selective antagonism of ionotropic glutamate receptors with kynurenic acid (2 nmol/50 nl) in the cNTS produced a significant increase in the baseline fR but no changes in the tachypneic response to chemoreflex activation. The selective antagonism of purinergic P2 receptors by PPADS (0.25 nmol/50 nl) in the cNTS produced no changes in the baseline fR or in the tachypneic response to chemoreflex activation. The data indicate that glutamate acting on ionotropic receptors in the cNTS plays a inhibitory role on the modulation of the baseline fR but had no effect on the tachypneic response to chemoreflex activation, while ATP acting on P2 receptors in the cNTS plays no major role in the modulation of the baseline fR or in the tachypneic response to chemoreflex activation. We suggest that neurotransmitters other than L-glutamate and ATP are involved in the processing of the tachypneic response of the chemoreflex at the cNTS level. (C) 2008 Elsevier B.V. All rights reserved.

Antifungal Resistance Mechanisms in Dermatophytes

Although fungi do not cause outbreaks or pandemics, the incidence of severe systemic fungal infections has increased significantly, mainly because of the explosive growth in the number of patients with compromised immune system. Thus, drug resistance in pathogenic fungi, including dermatophytes, is gaining importance. The molecular aspects involved in the resistance of dermatophytes to marketed antifungals and other cytotoxic drugs, such as modifications of target enzymes, over-expression of genes encoding ATP-binding cassette (ABC) transporters and stress-response-related proteins are reviewed. Emphasis is placed on the mechanisms used by dermatophytes to overcome the inhibitory action of terbinafine and survival in the host environment. The relevance of identifying new molecular targets, of expanding the understanding about the molecular mechanisms of resistance and of using this information to design new drugs or to modify those that have become ineffective is also discussed.

Interaction of purinergic and nitrergic mechanisms in the caudal nucleus tractus solitarii of rats

The interaction of purinergic and nitrergic mechanisms was evaluated in the caudal nucleus tractus solitarii (cNTS) using awake animals and brainstem slices. In awake animals, ATP (1.25 nmol/50 nL) was microinjected into the cNTS before and after the microinjection of a selective neuronal nitric oxide synthase (nNOS) inhibitor N-propyl-L-arginine (NPLA, 3 pmoles/50 nL, n=8) or vehicle (saline, n=4), and cardiovascular and ventilatory parameters were recorded. In brainstem slices from a distinct group of rats, the effects of ATP on the NO concentration in the cNTS using the fluorescent dye DAF-2 DA were evaluated. For this purpose brainstem slices (150 pm) containing the cNTS were pre-incubated with ATP (500 mu M; n=8) before and during DAF-2 DA loading. Microinjection of ATP into the cNTS increases the arterial pressure (AP), respiratory frequency (f(R)) and minute ventilation (V(E)), which were significantly reduced by pretreatment with N-PLA, a selective nNOS inhibitor (AP: 39 +/- 3 vs 16 +/- 14 mm Hg; f(R): 75 +/- 14 vs 4 +/- 3 cpm; V(E): 909 159 vs 77 39 mL kg(-1) m(-1)). The effects of ATP in the cNTS were not affected by microinjection of saline. ATP significantly increased the NO fluorescence in the cNTS (62 +/- 7 vs 101 +/- 10 AU). The data show that in the cNTS: a) the NO production is increased by ATP; b) NO formation by nNOS is involved in the cardiovascular and ventilatory responses to microinjection of ATP. Taken together, these data suggest an interaction of purinergic and nitrergic mechanisms in the cNTS. (C) 2009 Elsevier B.V. All rights reserved.

Response of human alveolar bone-derived cells to a novel poly(vinylidene fluoride-trifluoroethylene)/barium titanate membrane

This study investigated the response of human alveolar bone-derived cells to a novel poly(vinylidene fluoride-trifluoroethylene)/barium titanate (P(VDF-TrFE)/BT) membrane. Osteoblastic cells were cultured in osteogenic conditions either on P(VDF-TrFE)/BT or polytetrafluoroethylene (PTFE) for up to 14 days. At 7 and 14 days, the mRNA expression of Runt-related transcription factor 2 (RUNX2), Type I collagen (COL I), Osteopontin (OPN), Alkaline phosphatase (ALP), Bone sialoprotein (BSP), and Osteocalcin (OC), key markers of the osteoblastic phenotype, and of Bcl2-associated X protein (Bax), B-cell CLL/lymphoma 2 (Bcl-2), and Survivin (SUR), associated with the control of the apoptotic cell death, was assayed by real-time PCR. In situ ALP activity was qualitatively evaluated by means of Fast red staining. Surface characterization was also qualitatively and quantitatively assayed in terms of topography, roughness, and wettability. Cells grown on P(VDF-TrFE)/BT exhibited a significantly higher mRNA expression for all markers compared to the ones on PTFE, except for Bcl-2, which was not detected for both groups. Additionally, Fast red staining was noticeably stronger in cultures on P(VDF-TrFE)/BT at 7 and 14 days. At micron-and submicron scale, SEM images and roughness analysis revealed that PTFE and P(VDF-TrFE)/BT exhibited a smooth topography and a similar roughness, respectively. PTFE membrane displayed higher contact angles compared with P(VDF-TrFE)/BT, as indicated by wettability assay. The novel P(VDF-TrFE)/BT membrane supports the acquisition of the osteoblastic phenotype in vitro, while up-regulating the expression of apoptotic markers. Further in vivo experiments should be carried out to confirm the capacity of P(VDF-TrFE)/BT membrane in promoting bone formation in guided bone regeneration.

Serotonergic mechanisms on breathing modulation in the rat locus coeruleus

The locus coeruleus (LC) is a noradrenergic nucleus that plays an important role in the ventilatory response to hypercapnia. This nucleus is densely innervated by serotonergic fibers and contains high density of serotonin (5-HT) receptors, including 5-HT(1A) and 5-HT(2). We assessed the possible modulation of respiratory response to hypercapnia by 5-HT, through 5-HT(1A) and 5-HT(2) receptors, in the LC. To this end, we determined the concentrations of 5-HT and its metabolite 5-hydroxyindole-3-acetic acid (5-HIAA) in the LC after hypercapnic exposure. Pulmonary ventilation (V(E), plethysmograph) was measured before and after unilateral microinjection (100 nL) of WAY-100635 (5-HT(1A) antagonist, 5.6 and 56 mM), 8-OHDPAT (5-HT(1A/7) agonist, 7 and 15 mM), Ketanserin (5-HT(2A) antagonist, 3.7 and 37 mM), or (+/-)-2,5-dimethoxy-4-iodoamphetaminehydrochloride (DOI; 5-HT(2A) agonist, 6.7 and 67 mM) into the LC, followed by a 60-min period of 7% CO(2) exposure. Hypercapnia increased 5-HTIAA levels and 5-HIAA/5-HT ratio within the LC. WAY-100635 and 8-OHDPAT intra-LC decreased the hypercapnic ventilatory response due to a lower tidal volume. Ketanserin increased CO(2) drive to breathing and DOI caused the opposite response, both acting on tidal volume. The current results provide evidence of increased 5-HT release during hypercapnia in the LC and that 5-HT presents an inhibitory modulation of the stimulatory role of LC on hypercapnic ventilatory response, acting through postsynaptic 5-HT(2A) receptors in this nucleus. In addition, hypercapnic responses seem to be also regulated by presynaptic 5-HT(1A) receptors in the LC.

NEONATAL HANDLING AND THE MATERNAL ODOR PREFERENCE IN RAT PUPS: INVOLVEMENT OF MONOAMINES AND CYCLIC AMP RESPONSE ELEMENT-BINDING PROTEIN PATHWAY IN THE OLFACTORY BULB

Early-life environmental events, such as the handling procedure, can induce long-lasting alterations upon several behavioral and neuroendocrine systems. However, the changes within the pups that could be causally related to the effects in adulthood are still poorly understood. In the present study, we analyzed the effects of neonatal handling on behavioral (maternal odor preference) and biochemical (cyclic AMP response element-binding protein (CREB) phosphorylation, noradrenaline (NA), and serotonin (5-HT) levels in the olfactory bulb (OB)) parameters in 7-day-old male and female rat pups. Repeated handling (RH) abolished preference for the maternal odor in female pups compared with nonhandled (NH) and the single-handled (SH) ones, while in RH males the preference was not different than NH and SH groups. In both male and female pups, RH decreased NA activity in the OB, but 5-HT activity increased only in males. Since preference for the maternal odor involves the synergic action of NA and 5-HT in the OB, the maintenance of the behavior in RH males could be related to the increased 5-HT activity, in spite of reduction in the NA activity in the OB. RH did not alter CREB phosphorylation in the OB of both male and females compared with NH pups. The repeated handling procedure can affect the behavior of rat pups in response to the maternal odor and biochemical parameters related to the olfactory learning mechanism. Sex differences were already detected in 7-day-old pups. Although the responsiveness of the hypothalamic-pituitary-adrenal axis to stressors is reduced in the neonatal period, environmental interventions may impact behavioral and biochemical mechanisms relevant to the animal at that early age. (C) 2009 IBRO. Published by Elsevier Ltd. All rights reserved.

Chronic ouabain treatment exacerbates blood pressure elevation in spontaneously hypertensive rats: the role of vascular mechanisms

Objective Hypertensive rats are more sensitive to the pressor effects of acute ouabain than normotensive rats. We analyzed the effect of chronic ouabain (similar to 8.0 mu g/day, 5 weeks) treatment on the blood pressure of spontaneously hypertensive rats (SHRs) and Wistar-Kyoto rats and the contribution of vascular mechanisms. Methods Responses to acetylcholine and phenylephrine were analyzed in isolated tail arteries. Protein expression of endothelial nitric oxide synthase and cyclooxygenase-2 (COX-2) were also investigated. Results Ouabain treatment enhanced blood pressure only in SHRs. The pD(2) for acetylcholine was decreased in arteries from SHRs compared with Wistar-Kyoto rats, and ouabain did not change this parameter. However, ouabain was able to increase the pD(2) to phenylephrine in SHRs. Nitric oxide synthase inhibition with N(G)-nitro-L-arginine methyl ester or potassium channel blockade by tetraetylamonium increased the response to phenylephrine in SHRs, with a smaller increase in response observed in ouabain-treated SHRs. In addition, indomethacin (a COX inhibitor) and ridogrel (a thromboxane A(2) synthase inhibitor and prostaglandin H(2)/thromboxane A(2) receptor antagonist) decreased contraction to phenylephrine in tail rings from ouabain-treated SHRs. Protein expression of endothelial nitric oxide synthase was unaltered following ouabain treatment in SHRs, whereas COX-2 expression was increased. Conclusion Chronic ouabain treatment further increases the raised blood pressure of SHRs. This appears to involve a vascular mechanism, related to a reduced vasodilator influence of nitric oxide and endothelium-derived hyperpolarizing factor and increased production of vasoconstrictor prostanoids by COX-2. These data suggest that the increased plasma levels of ouabain could play an important role in the maintenance of hypertension and the impairment of endothelial function. J Hypertens 27:1233-1242 (C) 2009 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.