No abstract

Upon activation, macrophages undergo extensive metabolic rewiring 1 , 2 . Production of itaconate through the inducible enzyme IRG1 is a key hallmark of this process 3 . Itaconate inhibits succinate dehydrogenase (SDH) 4 , 5 , has electrophilic properties 6 , and is associated with a change in cytokine production 4 . Here, we compare the metabolic, electrophilic, and immunologic profiles of macrophages treated with unmodified itaconate and a panel of commonly used itaconate derivatives to examine its role. Using wild type and Irg1 −/− macrophages, we show that neither dimethyl itaconate (DI), 4-octyl itaconate (4OI), nor 4-monoethyl itaconate (4EI) are converted into intracellular itaconate, while exogenous itaconic acid readily enters macrophages. We find that only DI and 4OI induce a strong electrophilic stress response, in contrast to itaconate and 4EI. This correlates with their immunosuppressive phenotype: DI and 4OI inhibit IκBζ and pro-IL-1β induction, as well as IL-6, IL-10, and IFN-β secretion in an Nrf2-independent manner. In contrast, itaconate treatment only suppressed IL-1β secretion but not pro-IL-1β levels, and, surprisingly, strongly enhanced LPS-induced IFN-β secretion. Consistently, Irg1 −/− macrophages produced lower levels of interferon and reduced transcriptional activation of this pathway. Our work establishes itaconate as an immunoregulatory, rather than strictly immunosuppressive metabolite, and highlights the importance of using unmodified itaconate in future studies.

SUMMARY Itaconate is a unique regulatory metabolite that is induced upon Toll-like receptor (TLR) stimulation in myeloid cells. Here, we demonstrate major inflammatory tolerance and cell death phenotypes associated with itaconate production in activated macrophages. We show that endogenous itaconate is a key regulator of the signal 2 of NLR family pyrin domain containing 3 (NLRP3) inflammasome activation after long lipopolysaccharide (LPS) priming, which establishes tolerance to late NLRP3 inflammasome activation. We show that itaconate acts synergistically with inducible nitric oxide synthase (iNOS) and that the ability of various TLR ligands to establish NLRP3 inflammasome tolerance depends on the pattern of co-expression of IRG1 and iNOS. Mechanistically, itaconate accumulation upon prolonged inflammatory stimulation prevents full caspase-1 activation and processing of gasdermin D, which we demonstrate to be post-translationally modified by endogenous itaconate. Altogether, our data demonstrate that metabolic rewiring in inflammatory macrophages establishes tolerance to NLRP3 inflammasome activation that, if uncontrolled, can result in pyroptotic cell death and tissue damage.

No abstract

In the present study, mouse embryonic stem cells (ESCs) were differentiated into alveolar epithelial type II (AEII) cells for endotracheal injection. These enriched lung-like populations expressed lung epithelial markers SP-A, SP-B, SP-C, and CC10. First we show that rapid differentiation of ESCs requires a dissociated seeding method instead of an embryoid body culture method. We then investigated a two-step differentiation of ESCs into definitive endoderm by activin or A549-conditioned medium as a precursor to lung epithelial cells. When conditioned medium from A549 cells was used to derive endoderm, yield was increased above that of activin alone. Further studies showed that Wnt3a may be one of the secreted factors produced by A549 cells and promotes definitive endoderm differentiation, in part, through suppression of primitive endoderm. Activin and Wnt3a together at appropriate doses with dissociated cell seeding promoted greater endoderm yield than activin alone. Next, fibroblast growth factor 2 was shown to induce a dose-dependent expression of SPC, and these cells contained lamellar bodies characteristic of mature AEII cells from ESC-derived endoderm. Finally, ES-derived lung cells were endotracheally injected into preterm mice with evidence of AEII distribution within the lung parenchyma. This study concludes that a recapitulation of development may enhance derivation of an enriched population of lung-like cells for use in cell-based therapy.

Highlights d Combination checkpoint blockade leads to impaired efficacy with low tumor burden d This impairment results from IFN-g-mediated deletion of tumor-reactive T cells d AICD is an immune-intrinsic mechanism of therapeutic resistance to checkpoint blockade

Polypropylene ether glycols are used commercially as intermediates for flexible polyurethane foams and elastomers. In order to maximize polyurethane molecular weight and polymer properties concomitantly, highly bifunctional intermediates are required.Polypropylene ether glycols are prepared by the base-catalyzed polymerization of propylene oxide. A side reaction accompanying this polymerization gives rise to some monofunctioiiality by producing chains terminated with hydroxyl groups on one end but with ally1 or propenyl groups on the In addition, the glycols often contain an impurity detectable by its absorption in the carbonyl region of the infrared, and it was observed that the quality of polyurethanes prepared from such glycols varied inversely with the concentration of this impurity. It appeared, therefore, that the impurity represents a second type of monofunctionality.This study of the chemistry of propylene oxide polymerization was undertaken to gain some insight into the reactioiis leading t o the formation of carbonyl functions and unsaturation. EXPERIMENTALPropylene oxide was polymerized at atmospheric pressure using the apparatus shown in Figure 1. The oxide was placed in dropping funnel B ; 1,2-propanediol (initiator) and potassium hydroxide (catalyst) were placed in the reaction vessel and solution effected by heating at 100°C. Nitrogen continually fed under the liquid surface provided adequate agitation for this operation. After the potassium hydroxide had dissolved, propylene oxide was added slowly. Temperature control was maintained by varying the extent of immersion of the reactor in the oil bath, which was heated to about 20°C. higher than the desired polymerization temperature. I n spite of the slow feed rate, some of the monomer distilled out of the reactor and collected in storage vessel A from which it was periodically returned to dropping funnel B. Adequate agitation of the polymerization mass was provided by the vaporization of unreacted monomer. During the later stages of the polymerization, a positive nitrogen pressure in B 303

+ regulatory T (Treg) cells are generated during thymocyte development and play a crucial role in preventing the immune system from attacking the body's cells and tissues. However, how the formation of these cells is directed by T-cell receptor (TCR) recognition of self-peptide:major histocompatibility complex (MHC) ligands remains poorly understood. We show that an agonist self-peptide with which a TCR is strongly reactive can induce a combination of thymocyte deletion and CD4+ Treg cell formation in vivo. A weakly cross-reactive partial agonist self-peptide could similarly induce thymocyte deletion, but failed to induce Treg cell formation. These studies indicate that CD4+ Treg cell formation can require highly stringent recognition of an agonist self-peptide by developing thymocytes. They also refine the "avidity" model of thymocyte selection by demonstrating that the quality of the signal mediated by agonist self-peptides, rather than the overall intensity of TCR signaling, can be a critical factor in directing autoreactive thymocytes to undergo CD4+ Treg cell formation and/or deletion during their development.affinity | immune regulation | specificity | tolerance | transgenic mice