RESUMEN
Soluble guanylate cyclase (sGC) catalyzes the conversion of guanosine triphosphate into cyclic guanosine-3',5'-monophosphate, a key second messenger in cell signaling and tissue homeostasis. It was recently demonstrated that sGC stimulation is associated with a marked antiinflammatory effect in the liver of mice with experimental nonalcoholic steatohepatitis (NASH). Here, we investigated the mechanisms underlying the antiinflammatory effect of the sGC stimulator praliciguat (PRL) in the liver. Therapeutic administration of PRL exerted antiinflammatory and antifibrotic actions in mice with choline-deficient l-amino acid-defined high-fat diet-induced NASH. The PRL antiinflammatory effect was associated with lower F4/80- and CX3CR1-positive macrophage infiltration into the liver in parallel with lower Ly6CHigh- and higher Ly6CLow-expressing monocytes in peripheral circulation. The PRL antiinflammatory effect was also associated with suppression of hepatic levels of interleukin (IL)-1ß, NLPR3 (NACHT, LRR, and PYD domain-containing protein 3), ASC (apoptosis-associated speck-like protein containing a caspase-recruitment domain), and active cleaved-caspase-1, which are components of the NLRP3 inflammasome. In Kupffer cells challenged with the classical inflammasome model of lipopolysaccharide plus adenosine triphosphate, PRL inhibited the priming (expression of Il1b and Nlrp3) and blocked the release of mature IL-1ß. Mechanistically, PRL induced the protein kinase G (PKG)-mediated phosphorylation of the VASP (vasodilator-stimulated phosphoprotein) Ser239 residue which, in turn, reduced nuclear factor-κB (NF-κB) activity and Il1b and Nlrp3 gene transcription. PRL also reduced active cleaved-caspase-1 levels independent of pannexin-1 activity. These data indicate that sGC stimulation with PRL exerts antiinflammatory actions in the liver through mechanisms related to a PKG/VASP/NF-κB/NLRP3 inflammasome circuit.
Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Inflamasomas/metabolismo , Hígado/metabolismo , Proteínas de Microfilamentos/metabolismo , FN-kappa B/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Fosfoproteínas/metabolismo , Guanilil Ciclasa Soluble/metabolismo , Animales , Antiinflamatorios/metabolismo , Antiinflamatorios/farmacología , Antígenos Ly/metabolismo , Receptor 1 de Quimiocinas CX3C/metabolismo , Caspasa 1/metabolismo , Interleucina-1beta/metabolismo , Macrófagos del Hígado/metabolismo , Lipopolisacáridos , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Guanilil Ciclasa Soluble/farmacologíaRESUMEN
BACKGROUND AND AIM: Extracellular matrix (ECM) components released during excessive fat mass expansion are considered potential endogenous danger/alarm signals contributing to innate immune system activation. The aim of the current study was to specifically measure plasma levels of low molecular weight (LMW) hyaluronan (HA) and to evaluate its role as pro-inflammatory damage-associated molecular pattern (DAMP) on leukocyte response in the context of human obesity. SUBJECTS AND METHODS: Participants were selected according to their body mass index (BMI, kg/m2) as non-obese (BMI < 29.9, n = 18) and obese (BMI > 29.9, n = 33). Plasma samples were size-dependent fractionated using ion-exchange chromatography to specifically obtain LMW HA fractions that were subsequently quantified by ELISA. Cell incubation experiments with synthetic HA molecules were performed on freshly Ficoll-isolated neutrophils (PMN) and peripheral blood monocytes (PBMC). Leukocyte and adipose tissue gene expression was assessed by real-time PCR and NF-κB activation by western blot. Plasma cytokine levels were measured by fluorescent bead-based (Luminex) immunoassay. RESULTS: We observed a statistically significant increase in the circulating levels of HA fragments of LMW in individuals with obesity which were consistent with significant up-regulated expression of the LMW HA synthesizing enzyme hyaluronan synthase-1 (HAS-1) in obese adipose tissue. Gene expression assessment of HA receptors revealed up-regulated levels for TLR2 in both obese PMN and PBMC. Synthetic HA molecules of different sizes were tested on leukocytes from healthy donors. LMW HA fragments (15-40 kDa) and not those from intermediate molecular sizes (75-350 kDa) induced a significant up-regulation of the expression of major pro-inflammatory cytokines such as IL-1ß, MCP-1 and IL-8 in PBMC. Importantly, LMW HA was able to induce the phosphorylation of IKK α/ß complex supporting its pro-inflammatory role through NF-κB activation. CONCLUSION: Circulating LMW HA molecules are elevated in obesity and may play an important role in triggering low-grade inflammation and the development of metabolic complications.
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Ácido Hialurónico , Receptor Toll-Like 2 , Humanos , Ácido Hialurónico/química , Ácido Hialurónico/metabolismo , Ácido Hialurónico/farmacología , Receptor Toll-Like 2/metabolismo , FN-kappa B , Interleucina-8 , Leucocitos Mononucleares , Hialuronano Sintasas , Quinasa I-kappa B , Ficoll , Inflamación/metabolismo , Citocinas/metabolismo , Inmunidad Innata , ObesidadRESUMEN
Besides its oncotic power, albumin exerts pleiotropic actions, including binding, transport, and detoxification of endogenous and exogenous molecules, antioxidant activity, and modulation of immune and inflammatory responses. In particular, recent studies have demonstrated that albumin reduces leukocyte cytokine production. Here, we investigated whether albumin also has the ability to protect tissues from the damaging actions of these inflammatory mediators. We circumscribed our investigation to tumor necrosis factor (TNF) α, which exemplifies the connection between immunity and tissue injury. In vivo experiments in analbuminemic mice showed that these mice exhibit a more pronounced response to a model of TNFα-mediated liver injury induced by the administration of lipopolysaccharide (LPS) and D-galactosamine (D-gal). A tissue protective action against LPS/D-gal liver injury was also observed during the administration of human albumin to humanized mice expressing the human genes for albumin and neonatal Fc receptor (hAlb+/+ /hFcRn+/+ ) with preestablished carbon tetrachloride (CCl4 )-induced early cirrhosis. The cytoprotective actions of albumin against TNFα-induced injury were confirmed ex vivo, in precision-cut liver slices, and in vitro, in primary hepatocytes in culture. Albumin protective actions were independent of its scavenging properties and were reproduced by recombinant human albumin expressed in Oryza sativa. Albumin cytoprotection against TNFα injury was related to inhibition of lysosomal cathepsin B leakage accompanied by reductions in mitochondrial cytochrome c release and caspase-3 activity. These data provide evidence that in addition to reducing cytokines, the albumin molecule also has the ability to protect tissues against inflammatory injury.
Asunto(s)
Albúminas/metabolismo , Antiinflamatorios/farmacología , Hepatocitos/metabolismo , Cirrosis Hepática/metabolismo , Factor de Necrosis Tumoral alfa/toxicidad , Albúminas/farmacología , Albúminas/uso terapéutico , Animales , Antiinflamatorios/uso terapéutico , Tetracloruro de Carbono/toxicidad , Células Cultivadas , Hepatocitos/efectos de los fármacos , Hepatocitos/patología , Lipopolisacáridos/toxicidad , Cirrosis Hepática/tratamiento farmacológico , Cirrosis Hepática/etiología , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
OBJECTIVE: Systemic inflammation predisposes acutely decompensated (AD) cirrhosis to the development of acute-on-chronic liver failure (ACLF). Supportive treatment can improve AD patients, becoming recompensated. Little is known about the outcome of patients recompensated after AD. We hypothesise that different inflammasome activation is involved in ACLF development in compensated and recompensated patients. DESIGN: 249 patients with cirrhosis, divided into compensated and recompensated (previous AD), were followed prospectively for fatal ACLF development. Two external cohorts (n=327) (recompensation, AD and ACLF) were included. Inflammasome-driving interleukins (ILs), IL-1α (caspase-4/11-dependent) and IL-1ß (caspase-1-dependent), were measured. In rats, bile duct ligation-induced cirrhosis and lipopolysaccharide exposition were used to induce AD and subsequent recompensation. IL-1α and IL-1ß levels and upstream/downstream gene expression were measured. RESULTS: Patients developing ACLF showed higher baseline levels of ILs. Recompensated patients and patients with detectable ILs had higher rates of ACLF development than compensated patients. Baseline CLIF-C (European Foundation for the study of chronic liver failure consortium) AD, albumin and IL-1α were independent predictors of ACLF development in compensated and CLIF-C AD and IL-1ß in recompensated patients. Compensated rats showed higher IL-1α gene expression and recompensated rats higher IL-1ß levels with higher hepatic gene expression. Higher IL-1ß detection rates in recompensated patients developing ACLF and higher IL-1α and IL-1ß detection rates in patients with ACLF were confirmed in the two external cohorts. CONCLUSION: Previous AD is an important risk factor for fatal ACLF development and possibly linked with inflammasome activation. Animal models confirmed the results showing a link between ACLF development and IL-1α in compensated cirrhosis and IL-1ß in recompensated cirrhosis.
Asunto(s)
Insuficiencia Hepática Crónica Agudizada/etiología , Inflamasomas/efectos adversos , Cirrosis Hepática Experimental/complicaciones , Cirrosis Hepática/complicaciones , Adulto , Anciano , Anciano de 80 o más Años , Animales , Femenino , Humanos , Interleucina-1alfa/sangre , Interleucina-1alfa/metabolismo , Interleucina-1beta/sangre , Interleucina-1beta/metabolismo , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND & AIMS: Acute-on-chronic liver failure (ACLF) is a newly described syndrome, which develops in patients with acute decompensation of cirrhosis, and is characterized by intense systemic inflammation, multiple organ failures and high short-term mortality. The profile of circulating lipid mediators, which are endogenous signaling molecules that play a major role in inflammation and immunity, is poorly characterized in ACLF. METHODS: In the current study, we assessed the profile of lipid mediators by liquid chromatography coupled to tandem mass spectrometry in plasma from patients with acute decompensation of cirrhosis, with (n = 119) and without (n = 127) ACLF, and from healthy controls (n = 18). Measurements were prospectively repeated in 191 patients with acute decompensation of cirrhosis during a 28-day follow-up period. RESULTS: Fifty-nine lipid mediators (out of 100) were detected in plasma from cirrhotic patients, of which 16 were significantly associated with disease status. Among these, 11 lipid mediators distinguished patients at any stage from healthy controls, whereas 2 lipid mediators (LTE4 and 12-HHT, both derived from arachidonic acid) shaped a minimal plasma fingerprint that discriminated patients with ACLF from those without. Levels of LTE4 distinguished ACLF grade 3 from ACLF grades 1 and 2, followed the clinical course of the disease (increased with worsening and decreased with improvement) and positively correlated with markers of inflammation and non-apoptotic cell death. Moreover, LTE4 together with LXA5 (derived from eicosapentaenoic acid) and EKODE (derived from linoleic acid) were associated with short-term mortality. LXA5 and EKODE formed a signature associated with coagulation and liver failures. CONCLUSION: Taken together, these findings uncover specific lipid mediator profiles associated with disease severity and prognosis in patients with acute decompensation of cirrhosis. LAY SUMMARY: Acute-on-chronic liver failure (ACLF) is characterized by intense systemic inflammation, multiple organ failures and high short-term mortality. In the current study, we assessed the plasma lipid profile of 100 bioactive lipid mediators in healthy controls, patients with decompensated cirrhosis, and those who had developed ACLF. We identified lipid mediator signatures associated with inflammation and non-apoptotic cell death that discriminate disease severity and evolution, short-term mortality and organ failures.
Asunto(s)
Insuficiencia Hepática Crónica Agudizada/sangre , Metabolismo de los Lípidos , Lipidómica/métodos , Lípidos/sangre , Anciano , Biomarcadores/sangre , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Índice de Severidad de la Enfermedad , Factores de TiempoRESUMEN
Specialized proresolving mediators (SPMs) biosynthesized from docosahexaenoic acids (DHAs) including resolvins (Rvs), protectins, and maresins are potent endogenous autacoids that actively resolve inflammation, protect organs, and stimulate tissue regeneration. Our hypothesis was that failure of resolution programs may lead to unremitting inflammation in obesity, contributing to the development of metabolic comorbidities in this condition. Obese individuals with persistent low-grade systemic inflammation showed reduced leukocyte production of the DHA-derived monohydroxy fatty acid 17-hydroxy-DHA (HDHA) and unbalanced formation of SPMs (in particular D-series Rvs) accompanied by enhanced production of proinflammatory lipid mediators such as leukotriene B4. Mechanistic studies attributed this impairment to reduced 15-lipoxygenase (LOX) activity rather than altered DHA cellular uptake. Moreover, leukocytes from obese individuals exhibited decreased 5-LOX levels and reduced 5-LOX Ser271 phosphorylation and distinct intracellular 5-LOX redistribution. However, 15-LOX appears to be the most critical factor for the deficient production of SPMs by obese leukocytes because the formation of D-series Rvs was completely rescued by incubation with the intermediate precursor 17-HDHA. These data provide proof of concept that administration of intermediate precursors of SPM biosynthesis (e.g., 17-HDHA) could be more efficient in overriding impaired formation of these proresolving lipid mediators in conditions characterized by dysfunctional LOX activity, such as obesity.-López-Vicario, C., Titos, E., Walker, M. E., Alcaraz-Quiles, J., Casulleras, M., Durán-Güell, M., Flores-Costa, R., Pérez-Romero, N., Forné, M., Dalli, J., Clària, J. Leukocytes from obese individuals exhibit an impaired SPM signature.
Asunto(s)
Ácidos Docosahexaenoicos/biosíntesis , Leucocitos/metabolismo , Obesidad/metabolismo , Estudios de Casos y Controles , Ácidos Docosahexaenoicos/química , Humanos , Inflamación , Metabolismo de los LípidosRESUMEN
Decompensated cirrhosis is characterized by exuberant systemic inflammation. Although the inducers of this feature remain unknown, the presence of circulating forms of oxidized albumin, namely human nonmercaptalbumin 1 (HNA1) and HNA2, is a common finding in cirrhosis. The aim of this study was to explore the ability of these oxidized albumin forms to induce systemic inflammation by triggering the activation of peripheral leukocytes. We observed significantly higher plasma levels of HNA1 and HNA2 in patients with cirrhosis (n = 256) compared to healthy volunteers (n = 48), which gradually increased during the course from compensated to decompensated to acute-on-chronic liver failure. Plasma HNA1 and HNA2 levels significantly correlated with inflammatory markers (i.e., interleukin-6 [IL-6], IL-1ß, tumor necrosis factor-alpha [TNF-α] and IL-8) in patients with cirrhosis. To directly test the inflammatory effects of HNA1 and HNA2 on leukocytes, these oxidized albumin forms were prepared ex vivo and their posttranslational modifications monitored by liquid chromatography (LC)-quadrupole time-of-flight/mass spectrometry (MS). HNA1, but not HNA2, increased IL-1ß, IL-6, and TNF-α mRNA and protein expression in leukocytes from both healthy volunteers and patients with cirrhosis. Moreover, HNA1 up-regulated the expression of eicosanoid-generating enzymes (i.e., cyclooxygenase-2 [COX-2] and microsomal prostaglandin E [PGE] synthase 1) and the production of inflammatory eicosanoids (PGE2 , PGF2α , thromboxane B2 , and leukotriene B4 ), as determined by LC-electrospray ionization-MS/MS. The inflammatory response to HNA1 was more pronounced in peripheral blood mononuclear cells (PBMCs) and marginal in polymorphonuclear neutrophils. Kinome analysis of PBMCs revealed that HNA1 induced the phosphorylation of p38 mitogen-activated protein kinase, the inhibition of which blocked HNA1-induced cytokine and COX-2 induction. Conclusion: HNA1 triggers an inflammatory response in PBMCs, providing a rationale for its removal and replacement by reduced albumin in the prevention of systemic inflammation in patients with advanced liver disease.
Asunto(s)
Citocinas/metabolismo , Inflamación/metabolismo , Leucocitos/metabolismo , Cirrosis Hepática/metabolismo , Albúmina Sérica Humana/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Western Blotting , Cromatografía Liquida , Femenino , Humanos , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática/complicaciones , Fallo Hepático/etiología , Fallo Hepático/metabolismo , Masculino , Persona de Mediana Edad , Oxidación-Reducción , Especies Reactivas de Oxígeno/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Espectrometría de Masas en TándemRESUMEN
Acute-on-chronic liver failure (ACLF) in cirrhosis is an increasingly recognized syndrome characterized by acute decompensation, organ failure(s) and high short-term mortality. Recent findings suggest that an overexuberant systemic inflammation plays a primary role in ACLF progression. In this study, we examined whether genetic factors shape systemic immune responses in patients with decompensated cirrhosis. Six single-nucleotide polymorphisms (SNPs) in inflammation-related genes (interleukin [IL]-1 beta [IL-1ß], rs1143623; IL-1 receptor antagonist [IL-1ra], rs4251961; IL-10, rs1800871; suppressor of cytokine signaling-3, rs4969170; nucleotide-binding oligomerization domain-containing protein 2, rs3135500; and chemerin chemokine-like receptor 1, rs1878022) were genotyped in 279 patients with cirrhosis with (n = 178) and without (n = 101) ACLF from the CANONIC study of the CLIF consortium. Among these SNPs, we identified two polymorphisms belonging to the IL-1 gene cluster (IL-1ß and IL-1ra) in strong association with ACLF. Both SNPs were protective against ACLF; IL-1ß (odds ratio [OR], 0.34, 95% confidence interval [CI], 0.13-0.89; P < 0.05) and IL-1ra (OR, 0.58; 95% CI, 0.35-0.95; P < 0.05) under the recessive and overdominant inheritance models, respectively. These protective SNPs translated into reduced circulating levels of IL-1ß, IL-1α, IL-6, granulocyte-colony stimulating factor, granulocyte-macrophage colony-stimulating factor, and C-reactive protein at enrollment as well as after 7-14 days of admission. These findings were confirmed in vitro in leukocytes incubated with plasma from patients with decompensated cirrhosis carrying the protective SNP genotypes. Notably, a higher frequency of the protective genotypes was observed in patients without (80%) than in those with (20%) ACLF. Consistently, patients carrying the combined protective genotypes showed a lower 28-day mortality rate. CONCLUSION: These data identify two common functional polymorphisms in the IL-1 gene cluster, which are associated with the inflammatory process related to development of ACLF. (Hepatology 2017;65:202-216).
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Insuficiencia Hepática Crónica Agudizada/genética , Inflamación/genética , Interleucina-1/genética , Familia de Multigenes , Polimorfismo de Nucleótido Simple , Insuficiencia Hepática Crónica Agudizada/epidemiología , Femenino , Humanos , Inflamación/complicaciones , Cirrosis Hepática/complicaciones , Masculino , Persona de Mediana Edad , Factores de RiesgoRESUMEN
Endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) are hallmarks of nonalcoholic fatty liver disease (NAFLD), which is the hepatic manifestation of the metabolic syndrome associated with obesity. The specialized proresolving lipid mediator maresin 1 (MaR1) preserves tissue homeostasis by exerting cytoprotective actions, dampening inflammation, and expediting its timely resolution. Here, we explored whether MaR1 protects liver cells from lipotoxic and hypoxia-induced ER stress. Mice were rendered obese by high-fat diet feeding, and experiments were performed in primary hepatocytes, Kupffer cells, and precision-cut liver slices (PCLSs). Palmitate-induced lipotoxicity increased ER stress and altered autophagy in hepatocytes, effects that were prevented by MaR1. MaR1 protected hepatocytes against lipotoxicity-induced apoptosis by activating the UPR prosurvival mechanisms and preventing the excessive up-regulation of proapoptotic pathways. Protective MaR1 effects were also seen in hepatocytes challenged with hypoxia and TNF-α-induced cell death. High-throughput microRNA (miRNA) sequencing revealed that MaR1 actions were associated with specific miRNA signatures targeting both protein folding and apoptosis. MaR1 also prevented lipotoxic-triggered ER stress and hypoxia-induced inflammation in PCLSs and enhanced Kupffer cell phagocytic capacity. Together, these findings describe the ability of MaR1 to oppose ER stress in liver cells under conditions frequently encountered in NAFLD.-Rius, B., Duran-Güell, M., Flores-Costa, R., López-Vicario, C., Lopategi, A., Alcaraz-Quiles, J., Casulleras, M., Lozano, J. J., Titos, E., Clària, J. The specialized proresolving lipid mediator maresin 1 protects hepatocytes from lipotoxic and hypoxia-induced endoplasmic reticulum stress.
Asunto(s)
Antígenos Ly/metabolismo , Apoptosis/fisiología , Estrés del Retículo Endoplásmico/fisiología , Hepatocitos/metabolismo , Hipoxia/metabolismo , Receptor 1 Gatillante de la Citotoxidad Natural/metabolismo , Animales , Antígenos Ly/genética , Apoptosis/genética , Dieta Alta en Grasa/efectos adversos , Estrés del Retículo Endoplásmico/genética , Macrófagos del Hígado/metabolismo , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , MicroARNs/metabolismo , Receptor 1 Gatillante de la Citotoxidad Natural/genética , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Obesidad/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Persistent activation of the innate immune system greatly influences the risk for developing metabolic complications associated with obesity. In this study, we explored the therapeutic potential of the specialized proresolving mediator (SPM) resolvin D1 (RvD1) to actively promote the resolution of inflammation in human visceral adipose tissue from obese (Ob) patients. Using liquid chromatography-tandem mass spectrometry-based metabololipidomic analysis, we identified unbalanced production of SPMs (i.e., D- and E-series resolvins, protectin D1, maresin 1, and lipoxins) with respect to inflammatory lipid mediators (i.e., leukotriene B4 and PGs) in omental adipose tissue from Ob patients. In parallel, high-throughput transcriptomic analysis revealed a unique signature in this tissue that was characterized by overactivation of the IL-10 signaling pathway. Incubation of inflamed Ob visceral adipose tissues and human macrophages with RvD1 limited excessive activation of the IL-10 pathway by reducing phosphorylation of STAT proteins. Of interest, RvD1 blocked STAT-1 and its target inflammatory genes (i.e., CXCL9), as well as persistent STAT3 activation, without affecting the IL-10 anti-inflammatory response characterized by inhibition of IL-6, IL-1ß, IL-8, and TNF-α. Furthermore, RvD1 promoted resolution by enhancing expression of the IL-10 target gene heme oxygenase-1 by mechanisms dependent on p38 MAPK activity. Together, our data show that RvD1 can tailor the quantitative and qualitative responses of human inflamed adipose tissue to IL-10 and provide a mechanistic basis for the immunoresolving actions of RvD1 in this tissue. These findings may have potential therapeutic implications in obesity-related insulin resistance and other metabolic complications.
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Ácidos Docosahexaenoicos/inmunología , Inflamación/inmunología , Grasa Intraabdominal/inmunología , Transducción de Señal , Células Cultivadas , Ácidos Docosahexaenoicos/química , Humanos , Grasa Intraabdominal/patología , Obesidad/inmunología , Obesidad/patología , Obesidad/cirugíaRESUMEN
Soluble epoxide hydrolase (sEH) is an emerging therapeutic target in a number of diseases that have inflammation as a common underlying cause. sEH limits tissue levels of cytochrome P450 (CYP) epoxides derived from omega-6 and omega-3 polyunsaturated fatty acids (PUFA) by converting these antiinflammatory mediators into their less active diols. Here, we explored the metabolic effects of a sEH inhibitor (t-TUCB) in fat-1 mice with transgenic expression of an omega-3 desaturase capable of enriching tissues with endogenous omega-3 PUFA. These mice exhibited increased CYP1A1, CYP2E1, and CYP2U1 expression and abundant levels of the omega-3-derived epoxides 17,18-epoxyeicosatetraenoic acid (17,18-EEQ) and 19,20-epoxydocosapentaenoic (19,20-EDP) in insulin-sensitive tissues, especially liver, as determined by LC-ESI-MS/MS. In obese fat-1 mice, t-TUCB raised hepatic 17,18-EEQ and 19,20-EDP levels and reinforced the omega-3-dependent reduction observed in tissue inflammation and lipid peroxidation. t-TUCB also produced a more intense antisteatotic action in obese fat-1 mice, as revealed by magnetic resonance spectroscopy. Notably, t-TUCB skewed macrophage polarization toward an antiinflammatory M2 phenotype and expanded the interscapular brown adipose tissue volume. Moreover, t-TUCB restored hepatic levels of Atg12-Atg5 and LC3-II conjugates and reduced p62 expression, indicating up-regulation of hepatic autophagy. t-TUCB consistently reduced endoplasmic reticulum stress demonstrated by the attenuation of IRE-1α and eIF2α phosphorylation. These actions were recapitulated in vitro in palmitate-primed hepatocytes and adipocytes incubated with 19,20-EDP or 17,18-EEQ. Relatively similar but less pronounced actions were observed with the omega-6 epoxide, 14,15-EET, and nonoxidized DHA. Together, these findings identify omega-3 epoxides as important regulators of inflammation and autophagy in insulin-sensitive tissues and postulate sEH as a druggable target in metabolic diseases.
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Tejido Adiposo/metabolismo , Epóxido Hidrolasas/antagonistas & inhibidores , Ácidos Grasos Omega-3/metabolismo , Obesidad/metabolismo , Células 3T3-L1 , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/patología , Animales , Autofagia/fisiología , Benzoatos/farmacología , Cadherinas/genética , Cadherinas/metabolismo , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Inhibidores Enzimáticos/farmacología , Compuestos Epoxi/metabolismo , Ácido Graso Desaturasas/genética , Ácido Graso Desaturasas/metabolismo , Femenino , Inflamación/metabolismo , Inflamación/patología , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Ratones Transgénicos , Obesidad/patología , Compuestos de Fenilurea/farmacologíaRESUMEN
UNLABELLED: Acute-on-chronic liver failure (ACLF) in cirrhosis is characterized by acute decompensation (AD), organ failure(s), and high short-term mortality. Recently, we have proposed (systemic inflammation [SI] hypothesis) that ACLF is the expression of an acute exacerbation of the SI already present in decompensated cirrhosis. This study was aimed at testing this hypothesis and included 522 patients with decompensated cirrhosis (237 with ACLF) and 40 healthy subjects. SI was assessed by measuring 29 cytokines and the redox state of circulating albumin (HNA2), a marker of systemic oxidative stress. Systemic circulatory dysfunction (SCD) was estimated by plasma renin (PRC) and copeptin (PCC) concentrations. Measurements were performed at enrollment (baseline) in all patients and sequentially during hospitalization in 255. The main findings of this study were: (1) Patients with AD without ACLF showed very high baseline levels of inflammatory cytokines, HNA2, PRC, and PCC. Patients with ACLF showed significantly higher levels of these markers than those without ACLF; (2) different cytokine profiles were identified according to the type of ACLF precipitating event (active alcoholism/acute alcoholic hepatitis, bacterial infection, and others); (3) severity of SI and frequency and severity of ACLF at enrollment were strongly associated. The course of SI and the course of ACLF (improvement, no change, or worsening) during hospitalization and short-term mortality were also strongly associated; and (4) the strength of association of ACLF with SI was higher than with SCD. CONCLUSION: These data support SI as the primary driver of ACLF in cirrhosis. (Hepatology 2016;64:1249-1264).
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Insuficiencia Hepática Crónica Agudizada/complicaciones , Inflamación/etiología , Cirrosis Hepática/complicaciones , Insuficiencia Hepática Crónica Agudizada/sangre , Biomarcadores/sangre , Citocinas/sangre , Humanos , Inflamación/sangre , Cirrosis Hepática/sangreRESUMEN
Limited data regarding elevation of N-terminal pro-B-type natriuretic peptide (NT-proBNP) in mobilized donors with G-CSF is available. We extended these findings by examining serum NT-proBNP in a cohort study including 35 healthy donors and 69 patients who received G-CSF for CD34+ mobilization as well as 54 patients who did not receive G-CSF but who underwent collection of CD3+ cells for chimeric antigen receptor (CAR) T-cell manufacturing. No donor in the three cohorts experienced significant cardiac adverse events. NT-proBNP levels were measured before and after G-CSF administration and after finishing apheresis procedure. NT-proBNP increase was observed in mobilized healthy donors after G-CSF administration, but was not observed in mobilized or non-mobilized patients. Only in the cohort of healthy donors, pairwise comparisons using Wilcoxon signed ranks test showed a significant increase between the mean serum NT-proBNP level after G-CSF administration and the mean serum NT-proBNP level measured before G-CSF administration (231.09 ± 156.15 pg/mL vs. 58.88 ± 26.84 pg/mL; P < .01). No correlation was observed between NT-proBNP increase and G-CSF dose (rs = 0.09; n = 32; P = .6) and no other variables contributing to predict serum NT-proBNP increase were detected. In conclusion, we observed a statistically, although not clinically, significant increase of NT-proBNP in healthy donors who received G-CSF as CD34+ cell mobilization.
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Factor Estimulante de Colonias de Granulocitos , Movilización de Célula Madre Hematopoyética , Péptido Natriurético Encefálico , Fragmentos de Péptidos , Humanos , Péptido Natriurético Encefálico/sangre , Fragmentos de Péptidos/sangre , Masculino , Factor Estimulante de Colonias de Granulocitos/sangre , Femenino , Movilización de Célula Madre Hematopoyética/métodos , Persona de Mediana Edad , Adulto , Estudios de Cohortes , Anciano , Donantes de Sangre , Antígenos CD34RESUMEN
INTRODUCTION: The Spanish Haematology and Haemotherapy Society organizes peripheral blood smear review scheme, focused on the evaluation of diagnostic proficiency of participants by blood cell morphology analysis. The objective was to evaluate the efficacy of this scheme as an educational tool to improve the diagnostic proficiency of the participants. METHODS: During 2011-2019, 54 peripheral blood smears, alongside with patient details such as age, sex, blood cell counts and relevant clinical information, were sent to an average of 125 ± 13 laboratories per year. A number of 44 shipments were selected to analyse whether successive surveys of the same disease may lead to an improvement in the diagnostic success rate proposed by the laboratories. Participants were asked to select the most relevant morphological abnormalities, alongside the diagnostic orientation. Agreement of participant responses with RR was evaluated. RESULTS: Spanish laboratories showed a diagnostic proficiency greater than 80% in acute myeloid leukaemia, including acute promyelocytic leukaemia, mature B-cell neoplasms (hairy cell leukaemia and splenic marginal zone lymphoma), chronic myeloid leukaemia, sickle cell disease, Bernard-Soulier syndrome and infectious mononucleosis. It was important to note the significant improvement over the time in the successive shipments of the same disease, with a 31% and 13% increase in their diagnostic orientation success rate for acute myeloid leukaemia and acute promyelocytic leukaemia cases, respectively, 15% for mantle cell lymphoma and 6% for sickle cell disease. CONCLUSIONS: The present study provides evidence that peripheral blood smear review scheme can be a valid educational tool to improve the clinical pathologist skills in blood morphology and haematological diagnosis.
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Células Sanguíneas/patología , Neoplasias Hematológicas/sangre , Neoplasias Hematológicas/diagnóstico , Neoplasias Hematológicas/patología , Femenino , Humanos , Masculino , EspañaRESUMEN
Human serum albumin (HSA) is an emerging treatment for preventing excessive systemic inflammation and organ failure(s) in patients with acutely decompensated (AD) cirrhosis. Here, we investigated the molecular mechanisms underlying the immunomodulatory properties of HSA. Administration of HSA to patients with AD cirrhosis with elevated circulating bacterial DNA rich in unmethylated cytosine-phosphate-guanine dideoxynucleotide motifs (CpG-DNA) was associated with reduced plasma cytokine concentrations. In isolated leukocytes, HSA abolished CpG-DNA-induced cytokine expression and release independently of its oncotic and scavenging properties. Similar anti-inflammatory effects were observed with recombinant human albumin. HSA exerted widespread changes on the immune cell transcriptome, specifically in genes related to cytokines and type I interferon responses. Our data revealed that HSA was taken up by leukocytes and internalized in vesicles positively stained with early endosome antigen 1 and colocalized with CpG-DNA in endosomes, where the latter binds to Toll-like receptor 9 (TLR9), its cognate receptor. Furthermore, HSA also inhibited polyinosinic:polycytidylic acid- and lipopolysaccharide-induced interferon regulatory factor 3 phosphorylation and TIR domain-containing adapter-inducing interferon-ß-mediated responses, which are exclusive of endosomal TLR3 and TLR4 signaling, respectively. The immunomodulatory actions of HSA did not compromise leukocyte defensive mechanisms such as phagocytosis, efferocytosis, and intracellular reactive oxygen species production. The in vitro immunomodulatory effects of HSA were confirmed in vivo in analbuminemic humanized neonatal Fc receptor transgenic mice. These findings indicate that HSA internalizes in immune cells and modulates their responses through interaction with endosomal TLR signaling, thus providing a mechanism for the benefits of HSA infusions in patients with cirrhosis.
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Citocinas , Transducción de Señal , Albúminas , Humanos , Leucocitos , Cirrosis Hepática/tratamiento farmacológicoRESUMEN
The prototypic proinflammatory cytokine IL-1ß plays a central role in innate immunity and inflammatory disorders. The formation of mature IL-1ß from an inactive pro-IL-1ß precursor is produced via nonconventional multiprotein complexes called the inflammasomes, of which the most common is the nucleotide-binding domain leucine-rich repeat-containing protein 3 (NLRP3) inflammasome composed by NLRP3, (ASC) apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (CARD), and caspase-1. Specialized proresolving mediators (SPMs) promote resolution of inflammation, which is an essential process to maintain host health. SPMs prevent excessive inflammation by terminating the inflammatory response and returning to tissue homeostasis without immunosupression. This study tested the hypothesis that modulation of the NLRP3 inflammasome in macrophages is one mechanism involved in the SPM-regulated processes during resolution. Our findings demonstrate that the SPM resolvin D2 (RvD2) suppressed the expression of pro-IL-1ß and reduced the secretion of mature IL-1ß in bone marrow-derived macrophages challenged with LPS+ATP (classical NLRP3 inflammasome model) or LPS+palmitate (lipotoxic model). Similar findings were observed in thioglycolate-elicited peritoneal macrophages, in which RvD2 remarkably reduced ASC oligomerization, inflammasome assembly, and caspase-1 activity. In vivo, in a self-resolving zymosan A-induced peritonitis model, RvD2 blocked the NLRP3 inflammasome leading to reduced release of IL-1ß into the exudates, repression of osteopontin, and MCP-1 expression and induction of M2 markers of resolution (i.e., CD206 and arginase-1) in peritoneal macrophages. RvD2 inhibitory actions were receptor mediated and were abrogated by a selective GPR18 antagonist. Together, these findings support the hypothesis that SPMs have the ability to inhibit the priming and to expedite the deactivation of the NLRP3 inflammasome in macrophages during the resolution process.
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Ácidos Docosahexaenoicos/farmacología , Inflamasomas/metabolismo , Activación de Macrófagos , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Células de la Médula Ósea/citología , Caspasa 1/metabolismo , Inhibidores de Caspasas/farmacología , Inflamación/patología , Lipopolisacáridos , Activación de Macrófagos/efectos de los fármacos , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/metabolismo , Masculino , Ratones Endogámicos C57BL , Ácido Palmítico/farmacología , Fenotipo , ZimosanRESUMEN
Background: Patients with acutely decompensated cirrhosis (AD) may or may not develop acute-on-chronic liver failure (ACLF). ACLF is characterized by high-grade systemic inflammation, organ failures (OF) and high short-term mortality. Although patients with AD cirrhosis exhibit distinct clinical phenotypes at baseline, they have low short-term mortality, unless ACLF develops during follow-up. Because little is known about the association of profile of systemic inflammation with clinical phenotypes of patients with AD cirrhosis, we aimed to investigate a battery of markers of systemic inflammation in these patients. Methods: Upon hospital admission baseline plasma levels of 15 markers (cytokines, chemokines, and oxidized albumin) were measured in 40 healthy controls, 39 compensated cirrhosis, 342 AD cirrhosis, and 161 ACLF. According to EASL-CLIF criteria, AD cirrhosis was divided into three distinct clinical phenotypes (AD-1: Creatinine<1.5, no HE, no OF; AD-2: creatinine 1.5-2, and or HE grade I/II, no OF; AD-3: Creatinine<1.5, no HE, non-renal OF). Results: Most markers were slightly abnormal in compensated cirrhosis, but markedly increased in AD. Patients with ACLF exhibited the largest number of abnormal markers, indicating "full-blown" systemic inflammation (all markers). AD-patients exhibited distinct systemic inflammation profiles across three different clinical phenotypes. In each phenotype, activation of systemic inflammation was only partial (30% of the markers). Mortality related to each clinical AD-phenotype was significantly lower than mortality associated with ACLF (p < 0.0001 by gray test). Among AD-patients baseline systemic inflammation (especially IL-8, IL-6, IL-1ra, HNA2 independently associated) was more intense in those who had poor 28-day outcomes (ACLF, death) than those who did not experience these outcomes. Conclusions: Although AD-patients exhibit distinct profiles of systemic inflammation depending on their clinical phenotypes, all these patients have only partial activation of systemic inflammation. However, those with the most extended baseline systemic inflammation had the highest the risk of ACLF development and death.
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Insuficiencia Hepática Crónica Agudizada/patología , Inflamación/patología , Cirrosis Hepática/patología , Insuficiencia Hepática Crónica Agudizada/sangre , Anciano , Biomarcadores/sangre , Creatinina/sangre , Citocinas/sangre , Femenino , Humanos , Inflamación/sangre , Cirrosis Hepática/sangre , Masculino , Persona de Mediana Edad , Fenotipo , Pronóstico , Índice de Severidad de la EnfermedadRESUMEN
BACKGROUND AND PURPOSE: Non-alcoholic steatohepatitis (NASH) is the hepatic manifestation of metabolic syndrome and is characterized by steatosis, inflammation and fibrosis. Soluble guanylate cyclase (sGC) stimulation reduces inflammation and fibrosis in experimental models of lung, kidney and heart disease. Here, we tested whether sGC stimulation is also effective in experimental NASH. EXPERIMENTAL APPROACH: NASH was induced in mice by feeding a choline-deficient, l-amino acid-defined, high-fat diet. These mice received either placebo or the sGC stimulator IW-1973 at two different doses (1 and 3 mg·kg-1 ·day-1 ) for 9 weeks. IW-1973 was also tested in high-fat diet (HFD)-induced obese mice. Steatosis, inflammation and fibrosis were assessed by Oil Red O, haematoxylin-eosin, Masson's trichrome, Sirius Red, F4/80 and α-smooth muscle actin staining. mRNA expression was assessed by quantitative PCR. Levels of IW-1973, cytokines and cGMP were determined by LC-MS/MS, Luminex and enzyme immunoassay respectively. KEY RESULTS: Mice with NASH showed reduced cGMP levels and sGC expression, increased steatosis, inflammation, fibrosis, TNF-α and MCP-1 levels and up-regulated collagen types I α1 and α2, MMP2, TGF-ß1 and tissue metallopeptidase inhibitor 1 expression. IW-1973 restored hepatic cGMP levels and sGC expression resulting in a dose-dependent reduction of hepatic inflammation and fibrosis. IW-1973 levels were ≈40-fold higher in liver tissue than in plasma. IW-1973 also reduced hepatic steatosis and adipocyte hypertrophy secondary to enhanced autophagy in HFD-induced obese mice. CONCLUSIONS AND IMPLICATIONS: Our data indicate that sGC stimulation prevents hepatic steatosis, inflammation and fibrosis in experimental NASH. These findings warrant further evaluation of IW-1973 in the clinical setting.
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Inflamación/prevención & control , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Guanilil Ciclasa Soluble/efectos de los fármacos , Animales , Cromatografía Liquida/métodos , GMP Cíclico/metabolismo , Citocinas/metabolismo , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Cirrosis Hepática/prevención & control , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/patología , Reacción en Cadena de la Polimerasa , Guanilil Ciclasa Soluble/metabolismo , Espectrometría de Masas en Tándem/métodosRESUMEN
Obesity comorbidities are closely associated with chronic low-grade adipose tissue inflammation. A number of SNPs associated with inflammation has been identified, underscoring the impact of genetic determinants on this process. Here, we screened SNPs in genes with pro-inflammatory (IL-1ß, IL-6, STAT3 and JAK2), anti-inflammatory (IL-10 and SOCS3) and pro-resolving (ERV1/ChemR23) properties in 101 obese and 99 non-obese individuals. Among the SNPs analyzed, we identified that individuals carrying a C allele in the rs1878022 polymorphism of the ERV1/ChemR23 gene, which encodes for the receptor of the pro-resolving mediator RvE1, had increased ERV1/ChemR23 protein expression and reduced levels of the inflammatory cytokine IL-6 in adipose tissue. Moreover, patients carrying the C allele in homozygosity had lower plasma levels of IL-6, IFN-α2, IL-15, IL-1ra, IL-10, GM-CSF, G-CSF and VEGF and enhanced leukocyte responsiveness to RvE1. C-carriers also exhibited decreased TAG to HDL ratio, a surrogate marker of insulin resistance and a predictor of incident fatty liver. Finally, we confirmed in vivo that the ERV1/ChemR23 receptor regulates systemic and tissue inflammation since mice lacking ERV1/ChemR23 expression showed increased IL-6 levels in adipose tissue and peritoneal macrophages. Together, our study identified an ERV1/ChemR23 variant that protects patients with obesity from excessive inflammatory burden.
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Estudios de Asociación Genética , Inflamación/genética , Grasa Intraabdominal/patología , Obesidad Mórbida/genética , Polimorfismo de Nucleótido Simple/genética , Receptores de Quimiocina/genética , Animales , Femenino , Frecuencia de los Genes/genética , Homocigoto , Humanos , Patrón de Herencia/genética , Interleucina-6/metabolismo , Hígado/patología , Masculino , Ratones , Persona de Mediana Edad , Modelos Genéticos , Epiplón/patologíaRESUMEN
A novel genus of pro-resolving lipid mediators endogenously generated from omega-3 polyunsaturated fatty acids has been identified in exudates obtained during the resolution phase of acute inflammation. The term specialized pro-resolving mediators (SPM) has been coined for these lipid mediators, comprising four novel chemical mediator families designated resolvins of the E series (if derived from eicosapentaenoic acid) and resolvins of the D series, protectins and maresins (if generated from docosahexaenoic acid). These SPM act not only as "stop-signals" of inflammatory response, but also as facilitators of the ability of macrophages to clear apoptotic cells (efferocytosis) and migrate to peripheral lymph nodes (efflux), thus, expediting their removal from sites of inflammation. In this review, we provide an overview of the current efforts to elucidate the structure-function, biosynthesis and actions of these omega-3-derived SPM in the context of inflammatory diseases. We specifically highlight the role of these SPM as endogenous counter-regulators of the persistent inflammatory status present in adipose tissue of obese individuals and describe the potential therapeutic impact of these bioactive lipid autacoids on the prevention of hepatic co-morbidities associated with obesity and the metabolic syndrome.