The role of serum lipoxin A4 levels in the association between periodontal disease and metabolic syndrome

PURPOSE: An unresolved inflammatory state contributes to the pathogenesis of periodontal disease and metabolic syndrome (MetS). Therefore, the purpose of this study was to evaluate the role of lipoxin A4 (LXA4), a proresolving lipid mediator, in the association between periodontal disease and MetS. METHODS: Sixty-seven patients with MetS and 65 patients without MetS were included in the study. Sociodemographic information was obtained via a questionnaire, and detailed medical diagnoses were made. Periodontal parameters (plaque index [PI], gingival index [GI], probing pocket depth [PD], and clinical attachment level [CAL]) and metabolic parameters were measured, and serum LXA4 levels were determined. The associations among MetS, periodontal parameters, and serum LX levels were evaluated by adjusted multivariate linear regression analyses. RESULTS: Patients with MetS were older and had a higher body mass index than patients without MetS. Periodontal parameters (PI, GI, PD, and CAL) were higher in patients with MetS than in those without MetS. Serum LXA4 levels were higher in patients without MetS. Multivariate linear regression analysis indicated a positive association between MetS and periodontal parameters (PD and CAL). Negative associations were established between MetS and LXA4 levels, and between LXA4 and periodontal parameters (PI, PD, and CAL). CONCLUSIONS: The presence of higher values of periodontal parameters in patients with MetS and the negative relationship of LXA4 with MetS and periodontal disease may support the protective role of proresolving lipid mediators in the association between periodontal disease and MetS.

Complementary Participation of Genetics and Epigenetics in Development of NSAID-exacerbated Respiratory Disease

Nonsteroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (NERD) has attracted a great deal of attention because of its association with severe asthma. However, it remains widely underdiagnosed in asthmatics as well as the general population. Upon pharmacological inhibition of cyclooxygenase 1 by NSAIDs, production of anti-inflammatory prostaglandin E2 and lipoxins ceases, while release of proinflammatory cysteinyl leukotrienes increases. To determine the underlying mechanisms, many studies have attempted to elucidate the genetic variants, such as single nucleotide polymorphisms, responsible for alterations of prostaglandins and leukotrienes, but the results of these genetic studies could not explain the whole genetic pathogenesis of NERD. Accordingly, the field of epigenetics has been introduced as an additional contributor to genomic alteration underlying the development of NERD. Recently, changes in CpG methylation, as one of the epigenetic components, have been identified in target tissues of NERD. This review discusses in silico analyses of both genetic and epigenetic components to gain a better understanding of their complementary roles in the development of NERD. Although the molecular mechanisms underlying NERD pathogenesis remain poorly understood, genetic and epigenetic variations play significant roles. Our results enhance the understanding of the genetic and epigenetic mechanisms involved in the development of NERD and suggest new approaches toward better diagnosis and management.

Protective effect of early intervention with lipoxin A4 on septic mice / 中国当代儿科杂志

OBJECTIVE@#To study the effect of early intervention with lipoxin A4 (LXA4) on septic mice.@*METHODS@#Healthy male Balb/c mice aged 6-8 weeks were randomly divided into sham-operation group, sepsis group, 1-hour intervention group (intervention at 1 hour after sepsis), and 6-hour intervention group (intervention at 6 hours after sepsis) (n=8 each). A sepsis model was prepared by cecal ligation and puncture. The intervention groups received LXA4 at 0.01 μg/g body weight 1 or 6 hours after the model was established. Blood was taken from eyeballs at 24 hours after operation. Peritoneal lavage fluid and liver and lung tissue samples were collected. The bacterial colonies of whole blood and peritoneal lavage fluid were counted by dilution plating. The serum levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and monocyte chemoattractant protein-1 (MCP-1) were determined by cytometric bead array. The serum level of high mobility group box-1 (HGMB1) was determined using ELISA. The percentages of macrophages and neutrophils in peritoneal lavage fluid were determined by flow cytometry. Paraffin sectioning and hematoxylin-eosin staining were performed for the liver and lung tissue samples to observe pathological damage.@*RESULTS@#Compared with the sham-operation group, the sepsis group had a significantly decreased percentage of macrophages and a significantly increased percentage of neutrophils in peritoneal lavage fluid (P0.05). Compared with the 6-hour intervention group, the 1-hour intervention group had a significantly decreased serum level of HMGB1 (P0.05).@*CONCLUSIONS@#Early intervention with LXA4 may attenuate liver and lung injuries in septic mice, which may be explained by the decrease in serum levels of IL-6, TNF-α, MCP-1, and HMGB1, and it also may reduce the bacterial dissemination in the whole blood of septic mice, which may be explained by the increase in the percentage of peritoneal macrophages.

Effect of lipoxin A4 on the expression of Toll-like receptor 4 and TNF receptor-associated factor 6 in the liver of obese rats with sepsis / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the protective effect of lipoxin A4 (LXA4) against sepsis induced by lipopolysaccharide (LPS) in rats with obesity and its effect on the expression of Toll-like receptor 4 (TLR4) and TNF receptor-associated factor 6 (TRAF6) in the liver.</p><p><b>METHODS</b>A total of 60 male Sprague-Dawley rats aged three weeks were randomly divided into a normal group and an obesity group, with 30 rats in each group. A rat model of obesity was established by high-fat diet. Each of the two groups was further randomly divided into control group, sepsis group, and LXA4 group, and 8 rats were selected from each group. The rats in the control, sepsis, and LXA4 groups were treated with intraperitoneal injection of normal saline, LPS, and LXA4+LPS respectively. Twelve hours later, blood samples were collected from the heart and liver tissue samples were also collected. ELISA was used to measure the serum levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). Western blot was used to measure the protein expression of TLR4 and TRAF6 in liver tissue. Quantitative real-time PCR was used to measure the mRNA expression of TLR4 and TRAF6.</p><p><b>RESULTS</b>After being fed with high-fat diet for 6 weeks, the obesity group had significantly higher average weight and Lee's index than the normal group (P<0.05). Compared with the normal group, the obesity group had significant increases in the serum levels of IL-6 and TNF-α (P<0.05). In the normal group or the obesity group, the sepsis subgroup had significant increases in the serum levels of IL-6 and TNF-α compared with the control subgroup (P<0.05), while the LXA4 subgroup had significant reductions in the two indices compared with the sepsis subgroup (P<0.05). Compared with the normal group, the obesity group had significant increases in the protein and mRNA expression of TLR4 and TRAF6 (P<0.05). In the normal group or the obesity group, the sepsis subgroup had significant increases in the protein and mRNA expression of TLR4 and TRAF6 compared with the control subgroup (P<0.05). Compared with the sepsis subgroup, the LXA4 subgroup had significant reductions in the protein and mRNA expression of TLR4 and TRAF6 (P<0.05).</p><p><b>CONCLUSIONS</b>LXA4 can reduce the serum levels of IL-6 and TNF-α and alleviate inflammatory response. LXA4 can inhibit the expression of TLR4 and TRAF6 in the liver of septic rats, possibly by inhibiting the TLR4 signaling pathway.</p>

Lipoxin A4 Ameliorates Lipopolysaccharide-Induced A549 Cell Injury through Upregulation of N-myc Downstream-Regulated Gene-1 / 中华医学杂志(英文版)

<p><b>Background</b>Lipoxin A4 (LXA4) can alleviate lipopolysaccharide (LPS)-induced acute lung injury (ALI) and acute respiratory distress syndrome through promoting epithelial sodium channel (ENaC) expression in lung epithelial cells. However, how LXA4 promote ENaC expression is still largely elusive. The present study aimed to explore genes and signaling pathway involved in regulating ENaC expression induced by LXA4.</p><p><b>Methods</b>A549 cells were incubated with LPS and LXA4, or in combination, and analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) of ENaC-α/γ. Candidate genes affected by LXA4 were explored by transcriptome sequencing of A549 cells. The critical candidate gene was validated by qRT-PCR and Western blot analysis of A549 cells treated with LPS and LXA4 at different concentrations and time intervals. LXA4 receptor (ALX) inhibitor BOC-2 was used to test induction of candidate gene by LXA4. Candidate gene siRNA was adopted to analyze its influence on A549 viability and ENaC-α expression. Phosphoinositide 3-kinase (PI3K) inhibitor LY294002 was utilized to probe whether the PI3K signaling pathway was involved in LXA4 induction of candidate gene expression.</p><p><b>Results</b>The A549 cell models of ALI were constructed and subjected to transcriptome sequencing. Among candidate genes, N-myc downstream-regulated gene-1 (NDRG1) was validated by real-time-PCR and Western blot. NDRG1 mRNA was elevated in a dose-dependent manner of LXA4, whereas BOC-2 antagonized NDRG1 expression induced by LXA4. NDRG1 siRNA suppressed viability of LPS-treated A549 cells (treatment vs. control, 0.605 ± 0.063 vs. 0.878 ± 0.083, P = 0.040) and ENaC-α expression (treatment vs. control, 0.458 ± 0.038 vs. 0.711 ± 0.035, P = 0.008). LY294002 inhibited NDRG1 (treatment vs. control, 0.459 ± 0.023 vs. 0.726 ± 0.020, P = 0.001) and ENaC-α (treatment vs. control, 0.236 ± 0.021 vs. 0.814 ± 0.025, P < 0.001) expressions and serum- and glucocorticoid-inducible kinase 1 phosphorylation (treatment vs. control, 0.442 ± 0.024 vs. 1.046 ± 0.082, P = 0.002), indicating the PI3K signaling pathway was involved in regulating NDRG1 expression induced by LXA4.</p><p><b>Conclusion</b>Our research uncovered a critical role of NDRG1 in LXA4 alleviation of LPS-induced A549 cell injury through mediating PI3K signaling to restore ENaC expression.</p>

Remodelación vascular como modelo in vitro para el estudio del síndrome antifosfolipídico obstétrico / Vascular remodeling as in vitro model for the study of the obstetrical antiphospholipid syndrome

Introducción y Objetivo: Los anticuerpos antifosfolípidos (aAFL) se pueden unir a las células trofoblásticas o a las endoteliales, alterando la remodelación vascular y consecuentemente la placentación normal. El objetivo fue evaluar el efecto del suero de pacientes con síndrome antifosfolipídico (SAF) obstétrico en la interacción endotelio-trofoblasto utilizando un modelo in vitro tridimensional de remodelación vascular. Métodos: Las pacientes con aAFL fueron clasificadas en dos grupos: morbilidad gestacional y trombosis (MG/TV, n=7) y morbilidad gestacional sola (MG, n=8). Como control, se incluyeron mujeres sin aAFL con MG (MG/ aAFL-, n=10), y mujeres sanas (SHN, n=7). Células endoteliales HUVEC fueron cultivadas en Matrigel™ hasta formar estructuras tubulares (angiogénesis) y luego se adicionaron células trofoblásticas HTR8; estas células invaden las estructuras tubulares de las células endoteliales mejorando su estabilidad. Se evaluó el efecto de 10% del suero de las mujeres del estudio sobre esta interacción.

Effect of Alox-15 Polymorphism on GCF Levels of Lipoxin-A4 in Chronic Periodontitis: A Preliminary Study

Lipoxins play an important role in periodontal resolution, hence, investigation of genetic polymorphism of lipoxin gene may provide important information on the role of lipoxins in periodontal disease pathogenesis. The aim of this study was to investigate a polymorphism of C-to-T substitution at position c.-292 in ALOX15 (reticulocyte-type 15 lipoxygenase 1) gene in patients with chronic periodontitis and to associate the polymorphism with gingival crevicular fluid (GCF) lipoxin A4 (LXA4) levels. Forty-five chronic periodontitis and 45 periodontally healthy patients were included in this case-control study. Plaque index, calculus index, sulcus bleeding index, full mouth probing depth (PD) and clinical attachment loss (CAL) were recorded. GCF and blood samples were collected. GCF was analyzed for LXA4 levels by enzyme linked immunosorbant assay. Genotyping of ALOX15 polymorphism was studied using PCR. Mean LXA4 was lower in periodontitis group compared to the periodontally healthy group. There was a negative correlation between CAL and LXA4. The CC genotype was higher in the study group than in the control group. In the study group, mean CAL was significantly lower among individuals with the CT genotype. Mean LXA4 was significantly lower in CC genotype (45.0±7.11 ng/mL) compared to CT genotype (50.81±5.81 ng/mL) among the patients with periodontitis. The results suggest that LXA4 and c.-292T allele are associated with periodontal health. Polymorphisms in the ALOX15 gene may influence periodontal disease pathogenesis. Hence, investigation of such polymorphisms could benefit the evaluation of lipoxins role in periodontal disease.

Resumo Lipoxinas desempenham um papel importante na recuperação periodonta, portanto, a investigação do polimorfismo genético do gene da lipoxina pode fornecer informações importantes sobre o papel das lipoxinas na patogênese da doença periodontal. O objetivo deste estudo foi investigar um polimorfismo de substituição C-to-T na posição c-292 no gene ALOX15 (reticulócito-tipo 15 lipoxigenase 1) em pacientes com periodontite crônica e associar o polimorfismo com a lipoxina A4 (LXA4) do fluido gengival crevicular (FGC). Quarenta e cinco pacientes com periodontite crônica e 45 pacientes periodonalmente saudáveis foram incluídos neste estudo caso-controle. Índice de placa, índice de cálculo, índice de sangramento do sulco, profundidade de sondagem (PS) da boca toda e perda de inserção clínica (PIC) foram registrados. Amostras do FGC e de sangue foram coletadas. O FGC foi analisado quanto aos níveis de LXA4 por ensaio imunoadsorvente ligado à enzima (ELISA). A genotipagem do polimorfismo ALOX15 foi estudada por PCR. A média de LXA4 foi menor no grupo de periodontite em comparação com o grupo periodontalmente saudável. Houve uma correlação negativa entre PIC e LXA4. O genótipo CC foi maior no grupo de estudo do que no grupo controle. No grupo de estudo, a média de PIC foi significativamente menor entre os indivíduos com o genótipo CT. A média de LXA4 foi significativamente menor no genótipo CC (45,0 ± 7,11 ng / mL) em comparação com o genótipo CT (50,81 ± 5,81 ng / mL) entre os pacientes com periodontite. Os resultados sugerem que o alelo LXA4 e o alelo c-292T estão associados à saúde periodontal. Polimorfismos no gene ALOX15 podem influenciar a patogênese da doença periodontal. Assim, a investigação de tais polimorfismos pode beneficiar a avaliação do papel das lipoxinas na doença periodontal.

Suplementos dietarios en periodoncia: revisión bibliográfica narrativa / Additives in periodontics: a narrative review of the literature

Se realizó la revisión de los trabajos de investigación relacionados con la utilización de resolvinas y probióticos como complementos dietarios en la terapia periodontal, tanto en animales de experimentación como en humanos, describiendo el origen, la composición y los posibles mecanismos de acción. A partir de los trabajos revisados, se concluyó que las aludidas sustencias son, en la actualidad, un tema de investigación no traspolable a la clínica y que en el mediano plano, podrían potenciar las terapias actuales para patologías periodontales específicas.

Papel da Lipoxina A4 na fisiologia do sistema nervoso central e no dano cognitivo associado à neuroinflamação / Lipoxin A4 paper of the central nervous system physiology and cognitive impairment associated with neuroinflammation

O papel da lipoxina A4 (LXA4) no sistema imunológico é bem estudado, mas o papeldessa molécula na fisiologia do sistema nervoso central (SNC) só foi abordado recentemente.Como um modulador alostérico positivo, a LXA4 produz efeitos canabimiméticos e pode, dessaforma, estar envolvida em vßrios aspectos de funções fisiológicas reguladas pelo sistemaendocanabinóide (eCB). Nós investigamos essa hipótese analisando o comportamento decamundongos knockout (5-LO-/-) para a enzima 5-lipoxigenase (5-LO), que participa da síntesede LXA4. Ansiedade, consumo de ßgua e alimento, locomoção, nocicepção e memórias aversivassão comportamentos reconhecidos como sob controle do sistema eCB e foram avaliados nesteestudo. Nenhuma alteração foi observada no comportamento de 5-LO-/-em relação ao consumode ßgua e alimento e locomoção no teste de campo aberto. No entanto, o tratamento com LXA4produziu efeito ansiolítico no labirinto em cruz elevada. Além disso, inibição farmacológica da 5-LO demonstrou um efeito ansiogênico em animais idosos, mas não em adultos jovens, indicandoque a LXA4 endógena exerce um efeito regulatório sobre ansiedade de forma idade-dependente.Os animais 5-LO-/-apresentaram um aumento na sensibilidade e redução na tolerância à dor noteste de sensibilidade ao choque. Ainda, uma disfunção em memória de curto prazo e extinção dememória no teste de esquiva inibitória foi observada nos animais 5-LO-/-, indicando que a LXA4pode agir na facilitação do aprendizado...

The role of lipoxin A4 (LXA4) in the immune system is well studied, however the part thismolecule plays in central nervous system (CNS) physiology has only recently been addressed.LXA4, as a CB1 allosteric enhancer, has a cannabimimetic effect and may therefore be involvedin various aspects of endocannabinoid system (eCB) physiological functions. We investigatedthis reasoning by analyzing the behavior of 5-lipoxygenase (5-LO) knockout mice (5-LO-/-).Anxiety-like behavior, appetitive behavior, locomotion, nociception and learning and memory areall known to be under control of the eCB system and were assessed in this study. No alterationwas observed in the behavior of 5-LO knockout mice regarding appetitive behavior, measured byfood and water intake, or locomotion in the open field test. However, treatment with LXA4produced an anxiolytic-like effect on the elevated plus maze. Further, pharmacological inhibitionof 5-LO showed an anxiogenic-like effect in aged, but not adult mice, indicating that endogenousLXA4 has an age-dependent effect on the modulation of anxiety-like behavior. 5-LO-/- micepresented an increase in pain sensitivity and a decrease in pain tolerance in the foot shocksensitivity test. Interestingly, the animals also presented impairment in short-term memory andextinction learning in the step-down inhibitory avoidance task, pointing out that LXA4 may act inthe facilitation of fear learning. These data are of great importance to the possible use of LXA4 astreatment to neuroinflammation induced cognitive impairment.The study of sepsis reveals that an inflammatory stimulus can induce the exacerbatedproduction of pro-inflammatory cytokines in the CNS, which causes neuroinflammation andcognitive impairment...

Lipoxin A4 inhibits lipopolysaccharide-induced production of inflammatory cytokines in keratinocytes by up-regulating SOCS2 and down-regulating TRAF6 / 华中科技大学学报（医学）（英德文版）

Liopxin A4 (LXA4) is considered to be a crucial modulator in the inflammatory responses. In the present study, we aimed to study the effect of LXA4 on the inflammatory cytokines production induced by lipopolysaccharide (LPS) and the possible mechanism in normal human epidermal keratinocytes (NHEKs). NHEKs were isolated and cultured. The expression of toll-like receptor 4 (TLR4), LXA4 receptor (ALXR) and aryl hydrocarbon receptor (AhR) in NHEKs was detected by reverse transcription polymerase chain reaction (RT-PCR). The mRNA and protein levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1β (IL-1β) were determined in NHEKs stimulated by LPS (10 μg/mL) with or without preincubation with LXA4 (100 nmol/L) for 30 min by real-time quantitative PCR (real-time qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively. The expression levels of tumor necrosis factor receptor-associated factor 6 (TRAF6) and suppressors of cytokine signaling 2 (SOCS2) mRNAs and proteins, and nuclear translocation of NF-kB-p65 were measured by real-time qPCR and Western blotting, respectively. The results showed that NHEKs expressed TLR4, ALXR and AhR. LXA4 significantly inhibited the mRNA and protein expression levels of TNF-α, IL-1β and TRAF6 induced by LPS in NHEKs, and LXA4 obviously increased the expression of SOCS2 at mRNA and protein levels. The nuclear NF-kB-p65 protein expression induced by LPS was inhibited after preincubation with LXA4 in NHEKs. It was concluded that LXA4 inhibits the LPS-induced production of TNF-α and IL-1β in NHEKs by up-regulating SOCS2 and down-regulating TRAF6.

Change of serum myeloperoxidase and lipoxin A4 level in coronary heart disease patients with anxiety and/or depression / 中南大学学报(医学版)

OBJECTIVE@#To investigate the change of serum myeloperoxidase (MPO) and lipoxin A4 (LXA4) in coronary heart disease (CHD) patients with anxiety and depression and its clinical significance.@*METHODS@#From December 2010 to February 2011, 143 CHD patients and 44 non-CHD patients (the control group) hospitalized in the Department of Cardiology at the Second Xiangya Hospital were enrolled. The hospital anxiety and depression scale (HADS) was used to evaluate the psychological state of all patients and the CHD patients were assigned to an anxiety and depression group (n=57) or a non-depression and anxiety group (n=86). The serum levels of high sensitive C-reactive protein (Hs-CRP), MPO, and LXA4 were examined, and the ratio of MPO and LXA4 (M/L) was calculated.@*RESULTS@#The levels of Hs-CRP, MPO, and LXA4 as well as M/L ratios in both CHD groups were significantly higher than those in the control group (all P<0.01). Compared with the non-anxiety and depression group, the levels of MPO and LXA4, and M/L ratios in the anxiety and depression group increased (all P<0.05). Correlation analysis showed that MPO was positively correlated with the score of HADS-total (HADS-t), HADS-anxiety (HADS-a), or HADS-depression (HADS-d), while LXA4 was negatively correlated with HADS-t or HADS-d. Multiple ordinal logistic regression analysis revealed that higher HADS-t score, stable angina, unstable angina, and acute myocardial infarction were the independent impact factors for the elevation of M/L ratio.@*CONCLUSION@#Anxiety and depression may aggravate the inflammatory response in CHD patients. The imbalance between inflammation and anti-inflammation may be part of the mechanism.

Efeito do veneno de Crotalus durissus terrificus no edema crônico induzido na pata de camundongos pela injeção de bacilo Calmette-Guérin (BCG) / Effect of the Crotalus durissus terrificus in edema chronic induced in mice paw by the injection of bacilo Calmette-Guérin (BCG)

Diferentemente do observado em outros envenenamentos por serpentes da família Viperidae, nos causados pela Cdt não se observam sinais inflamatórios significativos no local da picada. Estudos prévios mostram que o VCdt inibe a resposta inflamatória aguda e algumas atividades biológicas de macrófagos, principal célula do processo inflamatório crônico. Nesse trabalho verificou-se o efeito do VCdt sobre o edema de pata crônico induzido pela injeção intraplantar de BCG em camundongos. Na concentração utilizada, o BCG evoca um edema crônico que é significativamente inibido pelo pré-tratamento dos animais com o VCdt (s.c.). Essa inibição persistiu por todo o período estudado (15 dias). O grupo que recebeu o veneno 1 h após a injeção de BCG também apresentou perfil de edema inibido em relação ao grupo controle, de forma semelhante ao observado no grupo pré-tratado com o veneno. Os grupos tratados com o VCdt 6 ou 11 dias após a injeção do BCG apresentavam edema de magnitude semelhante até o dia da injeção do veneno, sendo observada uma inibição significativa desse edema no dias subseqüentes à injeção do veneno. Uma vez constatado esse efeito inibitório do VCdt sobre o edema inflamatório crônico induzido pelo BCG, estudou-se qual a fração do veneno seria responsável por esse efeito. Os resultados indicam que a crotoxina, e não outros componentes deste veneno seja a responsável por essa inibição. Estudando-se possíveis mecanismos envolvidos nessa inibição, a reversão da inibição induzida pelo VCdt observada em grupos pré-tratados com dexametazona e zileuton, sugerem fortemente a participação de mediadores originados na via das lipoxigenases, no efeito inibitório do VCdt sobre esse modelo de inflamação crônica. Em conclusão, os resultados indicam que a crotoxina do VCdt possua uma significativa ação inibitória sobre o edema crônico induzido pelo BCG, possivelmente pela geração de mediadores antiinflamatórios da via das lipoxigenases.

Papel da endotelina-1 na inflamação articular / Role of endothelin-1 in the joint inflammation

As endotelinas (ET) são peptídeos produzidos por uma grande variedade de células e exercem seus efeitos através da sua ligação aos seus receptores de superfície ETA e ETB. Foi demonstrado que os níveis de ET-1 estão aumentados no plasma e no líquido sinovial de pacientes com artrite reumatóide (AR), uma doença autoimune crônica caracterizada por influxo celular, formação de edema e destruição articular. No entanto, apesar do aumento na produção de ET-1, o papel destes peptídeos durante a AR permanece pouco compreendido. O objetivo central deste estudo foi avaliar o papel das ET na migração celular e na formação de edema utilizando o modelo murino de artrite induzida por zymosan (AIZ). Nós observamos que a expressão de RNAm prepro-ET-1 encontra-se aumentada durante a AIZ e que o bloqueio dos receptores ETA e ETB reduziu significativamente a formação de edema, o recrutamento de neutrófilos para articulação e a produção de TNF-alfa, CXCL-1 e LTB4. Nós ainda demonstramos que o estímulo in vivo com ET-1 foi capaz de induzir a formação de edema, o recrutamento de neutrófilos e a produção de TNF-alfa, enquanto nossos dados in vitro demonstraram que a ET-1 é um potente estímulo quimiotático para neutrófilos. Em seguida, nós avaliamos os efeitos da lipoxina A4 (LXA4), um mediador lipídico pró-resolutivo e anti-inflamatório, durante a AIZ, focando nos efeitos da LXA4 sobre a expressão de ET-1 e sobre os efeitos próinflamatórios induzidos por ET-1 tanto in vivo quanto in vitro. O pré-tratamento com LXA4 reduziu a formação de edema, o influxo de neutrófilos, a expressão de RNAm para prepro-ET-1 e a produção articular de CXCL-1, LTB4 e TNF-alfa na articulação durante a AIZ. De acordo, o BML-111 (agonista do receptor LXA4) e o ácido acetil salicílico (AAS; responsável pela produção endógena de 15-epilipoxina) inibiram significativamente a formação de edema e o recrutamento de neutrófilos para a articulação estimulada com zymosan. Observamos também que o pré-tratamento in vivo com LXA4 durante a inflamação articular induzida por ET-1 inibiu o influxo celular, a formação de edema e a produção de TNF-alfa no lavado articular, enquanto o estímulo in vitro com LXA4 inibiu a ativação e a migração de neutrófilos induzida por ET-1. Resultados prévios obtidos durante a minha dissertação de mestrado demonstraram que uma fração enriquecida em tetranortriterpenóides obtida a partir da semente de Carapa guianensis Aublet apresentava um importante efeito anti-inflamatório na AIZ. No presente trabalho, nós demonstramos que a gedunina, uma das substâncias presentes na semente de C. guianensis, reduziu significativamente o influxo e a ativação celular induzidos por zymosan no líquido sinovial, que foi acompanhado pela diminuição da hipernocicepção e da produção de importantes mediadores inflamatórios envolvidos na AR, dentre eles a ET-1. Estudos in vitro demonstraram ainda que o tratamento com a gedunina foi capaz de impedir a ativação e a quimiotaxia de neutrófilos estimulados com ET-1. Tomados em conjunto, nossos dados demonstram que i) as ET exercem um papel pró-inflamatório central durante a AIZ, através da indução da produção de mediadores inflamatórios, formação de edema e migração de neutrófilos para as articulações inflamadas; ii) a LXA4 exerce importantes efeitos anti-inflamatórios durante a AIZ através da regulação da expressão e dos efeitos pró-inflamatórios in vitro e in vivo induzidos pela ET-1; iii) a gedunina exerce efeitos antiinflamatórios e anti-nociceptivos durante a AIZ e é capaz de inibir a expressão e a atividade de ET-1.

Effect of lipoxin A₄ on IL-1β production of monocytes and its possible mechanism in severe preeclampsia / 华中科技大学学报（医学）（英德文版）

This study examined in vitro effect of lipoxin A(4) (LXA(4)) on interleukin-1β (IL-1β) production of monocytes and its possible mechanism in severe preeclampsia (PE). Peripheral venous blood was drawn from 15 patients with severe preeclampsia (PE group) and 20 normal pregnant women (control group) to prepare monocytes which were then treated with LXA(4) at different concentrations of 0, 10, 100 nmol/L respectively. IL-1β level in the supernatant of monocytes was detected by enzyme linked immunoassay. The [Ca(2+)](i) of monocytes was measured by laser scanning confocal microscopy. The results showed that the IL-1β level and the [Ca(2+)](i) of monocytes in the PE group were significantly higher than those in the control group. LXA(4) significantly decreased the generation of IL-1β in a dose-dependent manner in the PE group. After treatment with 100-nmol/L LXA(4), in the PE group, the [Ca(2+)](i) concentration of monocytes was significantly reduced. It was concluded that LXA(4) may inhibit the IL-1β production of monocytes from severe preeclampsia women by inhibiting extracellular calcium influx.

Antemortem and postmortem estimation of skin wound age in rats: biochemical study

Inflammatory mediators play an important role in the mediation of inflammation and trauma. There for they could be usefulfor the determination of vitality and wound age. In the present study, interleukin-6 [IL-6], fibronectin [Fn] and lipoxinA[4] [LXA[4]] were estimated in extracts of antemortem skin wounds in rats. Moreover, we extended our measurements to include the levels of these mediators in rat skin wounds in the early postmortem period aiming to test for their practical usefulness in the estimation of wound vitality and the duration after its infliction. Thirty two rats were divided into 4 groups of rats [8 animals in each group] and were assigned for collection of wound samples at the indicated time intervals. The wound samples were taken 30 mm [group I], 3 hours [group II], 6 hours [group III] and 24 [group IV] hours after infliction of the incised wounds. The specimens as control group were excised from uninjured rats [8 animals] in the same region as wounded groufrs. The rats in group IV were sacrificed by cervical dislocation and kept at room temperature to be used in assessment of postmortem changes on different parameters. Postmortem wound samples were then collected 24 hours after sacrifice of group IV rats and the postmortem control samples were taken from intact skin of the same rats. A special group of rats [n-8] was used to explore the influence of supravital injuries on mediator release from postmortem inflicted wounds. All wound and control specimens were homogenated and assayed for the level of IL. 6, Fn and LXA[4] using quantitative ELISA analysis. Analysis of the changes of the studied parameters in different times revealed that Fn is the first mediator to increase, in 30 mm, after wound infliction. In the following 3 and 6 hours after wounding both Fn and IL-6 were increased. At 24 hours of wounding LXA[4] increases to join Fn and IL-6, at that time Fn and IL-6 were still high. This pattern of increased level of the three mediators was maintained for at least 24 hours postmortem. We can conclude that the combined assay of IL-6, Fn and LXA[4] may be a useful tool in determination of the probable duration lapsed since antemortem wound inflection. Moreover, this pattern of time dependent increase of the three parameters may be also useful in age determination of multiple inflicted wounds at variable intervals in the same victim. We can also conclude that vital reactions are essential for release of the assayed parameters. This can be documented by the lack of significant increase of these parameters in postmortem-inflicted wounds

Lipoxin A4 negatively regulates lipopolysaccharide-induced differentiation of RAW264.7 murine macrophages into dendritic-like cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Lipoxins (LXs), endogenous anti-inflammatory and pro-resolving eicosanoids generated during various inflammatory conditions, have novel immunomodulatory properties. Because dendritic cells (DCs) play crucial roles in the initiation and maintenance of immune response, we determined whether LXs could modulate the maturation process of DCs and investigated the effects of lipoxin A(4) (LXA(4)) on lipopolysaccharide (LPS)-induced differentiation of RAW264.7 cells into dendritic-like cells.</p><p><b>METHODS</b>RAW264.7 cells were cultured in vitro with 1 microg/ml LPS in the absence or presence of LXA(4) for 24 hours, and cellular surface markers (MHC-II, CD80 (B7-1), CD86 (B7-2)) were measured by flow cytometry (FCM). Mixed lymphocyte reaction was performed to evaluate the allostimulatory activity. Cytoplastic IkappaB degradation and nuclear factor kappa B (NF-kappaB) translocation were detected by Western blotting. Luciferase reporter plasmid was transiently transfected into RAW264.7 cells, and luciferase activity was determined to measure the transcriptional activity of NF-kappaB.</p><p><b>RESULTS</b>LXA(4) reduced the ratio of LPS-treated RAW264.7 cells to DCs with morphological characteristics and inhibited the expression of MHC II. LPS-induced up-regulation of CD86 was moderately suppressed by LXA(4) but no obvious change of CD80 was observed. Moreover, LXA(4) weakened the allostimulatory activity of LPS-treated RAW264.7 cells. These alterations of LPS+LXA(4)-treated cells were associated with a marked inhibition of IkappaB degradation, NF-kappaB translocation and then the transcriptional activity of NF-kappaB.</p><p><b>CONCLUSIONS</b>LXA(4) negatively regulates LPS-induced differentiation of RAW264.7 cells into dendritic-like cells. This activity reveals an undescribed mechanism of LXA(4) to prevent excessive and sustained immune reaction by regulating maturation of DCs.</p>

Mecanismos inatos de regulação da IL-12 durante a infecção por Mycobacterium tuberculosis / Innate mechanisms of regulation of IL-12 during Mycobacterium tuberculosis infection

A citocina IL-12 desempenha um papel importante na indução de uma rede de genes da resposta imune envolvidos na resistência a infecções por patógenos intracelulares. IL-12, citocina produzida principalmente por células dendríticas (DCs) e fagócitos, é uma das citocinas responsáveis pela ativação de uma resposta do tipo Thl, levando à produção de IFN-y e subsequente ativação de macrófagos, que se tornará um ambiente desfavorável para o sobrevivência de agentes invasores. Um exemplo desses microorganismos é o Mycobacterium tuberculosis, um importante patógeno humano causador da tuberculose. O M tuberculosis induz a produção de IL-12 em células apresentadoras de antígeno profissionais, entretanto, os mecanismos envolvidos na regulação dessa citocina durante o curso da infecção pelo bacilo não são complementamente entendidos. O objetivo geral desse projeto de tese é investigar os mecanismos imunes inatos que controlam a produção de IL-12 em resposta ao M tuberculosis. A hipótese testada foi que os receptores do tipo Tol! (TLR) e as lipoxinas são mediadores centrais da regulação de IL-12 influenciando a resistência/susceptibilidade ao bacilo. Um modelo experimental de infecção pulmonar e um modelo de infecção in vitro foram usados para esse propósito. amundongos deficientes em MyD88 se mostraram altamente susceptíveis à infecção via aerosol com M tuberculosis, implicando assim, a sinalização via TLR/IL-IR como um determinante da resposta do hospedeiro contra esse importante patógeno humano. Associado ao aumento de susceptibilidade, pulmões dos animais deficientes em MyD88 infectados com o bacilo apresentaram um significativa redução na indução de IL-12 e células T CD4+ secretoras de IFN-y. Observou-se também, que DCs de camundongos deficientes em MyD88 apresentaram uma falha na produção de IL-12 em resposta ao M tuberculosis in vitro. Apesar de trabalhos recentes indicarem que M tuberculosis contêm diversos ligantes como PIMs e DNA, os quais, podem...

Protective effects of 15-methyl-lipoxin A4 on mesangioproliferative nephritis in rats / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the protective effects of 15-methyl-lipoxin A4 (LXA4) on mesangioproliferative nephritis in rats and the possible mechanisms.</p><p><b>METHODS</b>Mesangioproliferative nephritis was induced by a single intravenous injection of the mouse monoclonal anti-Thy1.1 antibodies (ER4) in 20 rats. Ten nephritic rats were injected with 15-methyl-LXA4 at 10 minutes before ER4 antibody injection and then 8-hourly until the rats were sacrificed on day 4 after nephritis induction. The nephritis was evidenced by presence of proteinuria, histologic examination with light microscopy, infiltrating leukocyte assessed by immunofluorescence microscopy, and mesangial cell proliferation assessed by proliferation scoring and by immunohistochemical staining of proliferating cell nuclear antigen (PCNA). Expressions of interleukin (IL)-1beta and IL-6 protein or mRNA in glomeruli were determined by radioimmunoassay or RT-PCR, respectively. Phosphorylated phosphoinositide 3-kinase (PI3-K), Akt1 and p27(kip1) in glomeruli were analyzed by Western Blot. Activities of nuclear factor-kappaB (NF-kappaB) and signal transducer and activator of transcription 3 (STAT3) in glomeruli were assessed by electrophoretic mobility shift assay (EMSA).</p><p><b>RESULTS</b>There were increases in glomerular infiltration of leukocyte, expressions of IL-1beta and IL-6 protein and mRNA, and activities of NF-kappaB in nephritic rats between days 1 and 4 after nephritis induction. The enhanced proteinuria, score of mesangial proliferation, glomerular PCNA positive cells, activities of phosphorylated PI3-K, Akt1 and STAT3, and reduced p27(kip1) expression were found on day 4 after nephritis induction. 15-Methyl-LXA4 treatment significantly reduced the proteinuria, glomerular infiltration of leukocyte, expressions of IL-1beta and IL-6 protein and mRNA, score of mesangial proliferation, glomerular PCNA positive cells, activities of phosphorylated PI3-K, Akt1, NF-kappaB and STAT3, and increased the p27(kip1) expression.</p><p><b>CONCLUSIONS</b>15-Methyl-LXA4 can markedly inhibit the proteinuria, glomerular inflammation, and mesangial cell proliferation induced by anti-Thy1.1 antibodies. The inhibition effects are related to PI3-K/Akt1/p27(kip1)/cyclin pathway, STAT3 and NF-kappaB pathway-dependent signal transduction.</p>

New insights into the anti-inflammatory actions of aspirin- induction of nitric oxide through the generation of epi-lipoxins

Aspirin has always remained an enigmatic drug. Not only does it present with new benefits for treating an ever-expanding list of apparently unrelated diseases at an astounding rate but also because aspirin enhances our understanding of the nature of these diseases processe. Originally, the beneficial effects of aspirin were shown to stem from its inhibition of cyclooxygenase-derived prostaglandins, fatty acid metabolites that modulate host defense. However, in addition to inhibiting cyclooxygenase activity aspirin can also inhibit pro-inflammatory signaling pathways, gene expression and other factors distinct from eicosanoid biosynthesis that drive inflammation as well as enhance the synthesis of endogenous protective anti-inflammatory factors. Its true mechanism of action in anti-inflammation remains unclear. Here the data from a series of recent experiments proposing that one of aspirin's predominant roles in inflammation is the induction of nitric oxide, which potently inhibits leukocyte/endothelium interaction during acute inflammation, will be discussed. It will be argued that this nitric oxide-inducing effects are exclusive to aspirin due to its unique ability, among the family of traditional anti-inflammatory drugs, to acetylate the active site of inducible cyclooxygenase and generate a family of lipid mediators called the epi-lipoxins that are increasingly being shown to have profound roles in a range of host defense responses.

Lipoxin agonists: turn right! to path of resolving neutrophil

An impressive array of cellular and molecular adaptive responses achieves homeostasis. The inflammatory reaction is an adaptive response triggered by an insult to culminate into the overt cardinal signs of inflammation, eventually leading to resolution and returning the organism back to its original centered state. This article focuses on some aspects of the lipoxin A4 signaling pathway during the resolution phase, to better understand molecular mechanisms by which a neutrophil directs an inflammatory reaction to switch off and resume homeostasis. Defining the resolution state of a neutrophil at the molecular level will aid in treatments of diseases that are associated with an exaggerated and uncontrolled inflammation.