Anti-Inflammatory Activities of Inotilone from Phellinus linteus through the Inhibition of MMP-9, NF-κB, and MAPK Activation In Vitro and In Vivo

Inotilone was isolated from Phellinus linteus. The anti-inflammatory effects of inotilone were studied by using lipopolysaccharide (LPS)-stimulated mouse macrophage RAW264.7 cells and λ-carrageenan (Carr)-induced hind mouse paw edema model. Inotilone was tested for its ability to reduce nitric oxide (NO) production, and the inducible nitric oxide synthase (iNOS) expression. Inotilone was tested in the inhibitor of mitogen-activated protein kinase (MAPK) [extracellular signal-regulated protein kinase (ERK), c-Jun NH2-terminal kinase (JNK), p38], and nuclear factor-κB (NF-κB), matrix-metalloproteinase (MMP)-9 protein expressions in LPS-stimulated RAW264.7 cells. When RAW264.7 macrophages were treated with inotilone together with LPS, a significant concentration-dependent inhibition of NO production was detected. Western blotting revealed that inotilone blocked the protein expression of iNOS, NF-κB, and MMP-9 in LPS-stimulated RAW264.7 macrophages, significantly. Inotilone also inhibited LPS-induced ERK, JNK, and p38 phosphorylation. In in vivo tests, inotilone decreased the paw edema at the 4th and the 5th h after Carr administration, and it increased the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx). We also demonstrated that inotilone significantly attenuated the malondialdehyde (MDA) level in the edema paw at the 5th h after Carr injection. Inotilone decreased the NO and tumor necrosis factor (TNF-α) levels on serum at the 5th h after Carr injection. Western blotting revealed that inotilone decreased Carr-induced iNOS, cyclooxygenase-2 (COX-2), NF-κB, and MMP-9 expressions at the 5th h in the edema paw. An intraperitoneal (i.p.) injection treatment with inotilone diminished neutrophil infiltration into sites of inflammation, as did indomethacin (Indo). The anti-inflammatory activities of inotilone might be related to decrease the levels of MDA, iNOS, COX-2, NF-κB, and MMP-9 and increase the activities of CAT, SOD, and GPx in the paw edema through the suppression of TNF-α and NO. This study presents the potential utilization of inotilone, as a lead for the development of anti-inflammatory drugs

Glycoinositolphospholipids from Leishmania braziliensis and L. infantum: Modulation of Innate Immune System and Variations in Carbohydrate Structure

The essential role of the lipophosphoglycan (LPG) of Leishmania in innate immune response has been extensively reported. However, information about the role of the LPG-related glycoinositolphospholipids (GIPLs) is limited, especially with respect to the New World species of Leishmania. GIPLs are low molecular weight molecules covering the parasite surface and are similar to LPG in sharing a common lipid backbone and a glycan motif containing up to 7 sugars. Critical aspects of their structure and functions are still obscure in the interaction with the vertebrate host. In this study, we evaluated the role of those molecules in two medically important South American species Leishmania infantum and L. braziliensis, causative agents of visceral (VL) and cutaneous Leishmaniasis (CL), respectively. GIPLs derived from both species did not induce NO or TNF-α production by non-primed murine macrophages. Additionally, primed macrophages from mice (BALB/c, C57BL/6, TLR2−/− and TLR4−/−) exposed to GIPLs from both species, with exception to TNF-α, did not produce any of the cytokines analyzed (IL1-β, IL-2, IL-4, IL-5, IL-10, IL-12p40, IFN-γ) or p38 activation. GIPLs induced the production of TNF-α and NO by C57BL/6 mice, primarily via TLR4. Pre incubation of macrophages with GIPLs reduced significantly the amount of NO and IL-12 in the presence of IFN-γ or lipopolysaccharide (LPS), which was more pronounced with L. braziliensis GIPLs. This inhibition was reversed after PI-specific phospholipase C treatment. A structural analysis of the GIPLs showed that L. infantum has manose rich GIPLs, suggestive of type I and Hybrid GIPLs while L. braziliensis has galactose rich GIPLs, suggestive of Type II GIPLs. In conclusion, there are major differences in the structure and composition of GIPLs from L. braziliensis and L. infantum. Also, GIPLs are important inhibitory molecules during the interaction with macrophages

Neonatal and pediatric intensive care in Rio de Janeiro State, Brazil: an analysis of bed distribution, 1997 and 2007 Terapia intensiva neonatal e pediátrica no Rio de Janeiro, Brasil: distribuição de leitos e análise comparativa de equidade em 1997 e 2007

The objective of this study was to describe the characteristics of neonatal and pediatric intensive care units (ICU) and beds in Rio de Janeiro, correlating with population demands in 1997 and 2007. All neonatal and pediatric ICUs were visited, identifying the availability and type of beds. Comparisons were made between: supply and demand using projected need for beds for the population; public and private ICUs; and geographical regions. In 2007, 95 units were included totaling 1,094 beds (74 units and 1,080 beds in 1997): 51% public and 48% private (47% and 52% in 1997); 47% neonatal, 18% pediatric and 35% mixed units. Most units were located in the metropolitan area. The distribution of public and private beds was similar in the metropolitan area in both periods; in the interior, public beds tripled. Access has improved, mainly in the interior, but there is still no equity in the distribution of and accessibility to the available beds, with a shortage in the public sector, an excess in the private sector, and a great concentration in the metropolitan area.<br>Descrever as características das unidades de terapia intensiva (UTI) neonatais e pediátricas e leitos no Rio de Janeiro, Brasil, correlacionando com demandas da população em 1997 e 2007. UTIs neonatais e pediátricas foram visitadas, identificando-se o tipo e disponibilidade de leitos. Foram feitas comparações entre: a oferta e a demanda projetada da necessidade de leitos para a população, a natureza pública ou privada das UTIs e regiões geográficas. Em 2007, 95 unidades foram incluídas, totalizando 1.094 leitos (74 e 1.080 leitos em 1997): 51% públicas e 48% privadas (47% e 52% em 1997); 47% neonatais, 18% pediátricas e 35% mistas. A maioria estava localizada na região metropolitana. A distribuição dos leitos públicos e privados foi semelhante na região metropolitana em ambos os períodos, no interior os públicos triplicaram. O acesso melhorou, principalmente no interior, mas ainda não há equidade na distribuição e no acesso aos leitos disponíveis, com falta no setor público, excesso no privado, e grande concentração na região metropolitana

The curative effect of fucoidan on visceral leishmaniasis is mediated by activation of MAP kinases through specific protein kinase C isoforms

Fucoidan can cure both antimony-sensitive and antimony-resistant visceral leishmaniasis through immune activation. However, the signaling events underlying this cellular response remain uncharacterized. The present study reveals that fucoidan induces activation of p38 and ERK1/2 and NF-kB DNA binding in both normal and Leishmania donovani-infected macrophages, as revealed by western blotting and electrophoretic mobility shift assay (EMSA), respectively. Pharmacological inhibition of p38, ERK1/2 or the NF-kB pathway markedly attenuated fucoidan-induced pro-inflammatory cytokine synthesis and inducible nitric oxide synthase (iNOS) gene transcription, resulting in a reduction of parasite clearance. To decipher the underlying mechanism of fucoidan-mediated parasite suppression, the expression and functionality of various protein kinase C (PKC) isoforms were evaluated by immunoblotting and enzyme activity assay. Fucoidan elicited an increase in expression and activity of PKC-a, -bI and -bII isoforms in infected macrophages. Functional knockdown of PKC-a and -b resulted in downregulation of p38 and ERK1/2, along with a marked reduction of IL-12 and TNF-a production in fucoidan-treated infected macrophages. Collectively, these results suggest that the curative effect of fucoidan is mediated by PKC-dependent activation of the mitogen-activated protein kinase (MAPK)/NF-kB pathway, which ultimately results in the production of nitric oxide (NO) and disease-resolving pro-inflammatory cytokines