Increased Serum Levels of N-terminal pro-B-type Natriuretic Peptide (NT-proBNP) in Mobilized Healthy Donors with G-CSF: A Cohort Study.

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.

Albumin protects the liver from tumor necrosis factor α-induced immunopathology.

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.

Peripheral blood morphology review and diagnostic proficiency evaluation by a new Spanish EQAS during the period 2011-2019.

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.

Stimulation of soluble guanylate cyclase exerts antiinflammatory actions in the liver through a VASP/NF-κB/NLRP3 inflammasome circuit.

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.

Frontline Science: Specialized proresolving lipid mediators inhibit the priming and activation of the macrophage NLRP3 inflammasome.

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.

Oxidized Albumin Triggers a Cytokine Storm in Leukocytes Through P38 Mitogen-Activated Protein Kinase: Role in Systemic Inflammation in Decompensated Cirrhosis.

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.

Association of a variant in the gene encoding for ERV1/ChemR23 with reduced inflammation in visceral adipose tissue from morbidly obese individuals.

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.

The specialized proresolving lipid mediator maresin 1 protects hepatocytes from lipotoxic and hypoxia-induced endoplasmic reticulum stress.

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.

Prostaglandin E2 Exerts Multiple Regulatory Actions on Human Obese Adipose Tissue Remodeling, Inflammation, Adaptive Thermogenesis and Lipolysis.

Obesity induces white adipose tissue (WAT) dysfunction characterized by unremitting inflammation and fibrosis, impaired adaptive thermogenesis and increased lipolysis. Prostaglandins (PGs) are powerful lipid mediators that influence the homeostasis of several organs and tissues. The aim of the current study was to explore the regulatory actions of PGs in human omental WAT collected from obese patients undergoing laparoscopic bariatric surgery. In addition to adipocyte hypertrophy, obese WAT showed remarkable inflammation and total and pericellular fibrosis. In this tissue, a unique molecular signature characterized by altered expression of genes involved in inflammation, fibrosis and WAT browning was identified by microarray analysis. Targeted LC-MS/MS lipidomic analysis identified increased PGE2 levels in obese fat in the context of a remarkable COX-2 induction and in the absence of changes in the expression of terminal prostaglandin E synthases (i.e. mPGES-1, mPGES-2 and cPGES). IPA analysis established PGE2 as a common top regulator of the fibrogenic/inflammatory process present in this tissue. Exogenous addition of PGE2 significantly reduced the expression of fibrogenic genes in human WAT explants and significantly down-regulated Col1α1, Col1α2 and αSMA in differentiated 3T3 adipocytes exposed to TGF-ß. In addition, PGE2 inhibited the expression of inflammatory genes (i.e. IL-6 and MCP-1) in WAT explants as well as in adipocytes challenged with LPS. PGE2 anti-inflammatory actions were confirmed by microarray analysis of human pre-adipocytes incubated with this prostanoid. Moreover, PGE2 induced expression of brown markers (UCP1 and PRDM16) in WAT and adipocytes, but not in pre-adipocytes, suggesting that PGE2 might induce the trans-differentiation of adipocytes towards beige/brite cells. Finally, PGE2 inhibited isoproterenol-induced adipocyte lipolysis. Taken together, these findings identify PGE2 as a regulator of the complex network of interactions driving uncontrolled inflammation and fibrosis and impaired adaptive thermogenesis and lipolysis in human obese visceral WAT.