SignificanceAdipose tissue macrophages (ATMs) maintain adipose tissue homeostasis. However, during obesity ATMs become inflammatory, resulting in impaired adipose tissue function. Oxidative stress increases during obesity, which is thought to contribute to adipose tissue inflammation. To date, the connection between oxidative stress and adipose tissue inflammation remain unclear. In this study, we identify two classes of phospholipid oxidation products in lean and obese adipose tissue, which polarize macrophages to an antioxidant or proinflammatory state, respectively. Furthermore, we show that these phospholipids differently affect macrophage cellular metabolism, reflecting the metabolisms of ATMs found in lean and obese adipose tissue. Identification of pathways controlling ATM metabolism will lead to novel therapies for insulin resistance.
ObjectiveMacrophages control tissue homeostasis and inflammation by sensing and responding to environmental cues. However, the metabolic adaptation of macrophages to oxidative tissue damage and its translation into inflammatory mechanisms remains enigmatic.MethodsHere we identify the critical regulatory pathways that are induced by endogenous oxidation-derived DAMPs (oxidized phospholipids, OxPL) in vitro, leading to formation of a unique redox-regulatory metabolic phenotype (Mox), which is strikingly different from conventional classical or alternative macrophage activation.ResultsUnexpectedly, metabolomic analyses demonstrated that Mox heavily rely on glucose metabolism and the pentose phosphate pathway (PPP) to support GSH production and Nrf2-dependent antioxidant gene expression. While the metabolic adaptation of macrophages to OxPL involved transient suppression of aerobic glycolysis, it also led to upregulation of inflammatory gene expression. In contrast to classically activated (M1) macrophages, Hif1α mediated expression of OxPL-induced Glut1 and VEGF but was dispensable for Il1β expression. Mechanistically, we show that OxPL suppress mitochondrial respiration via TLR2-dependent ceramide production, redirecting TCA metabolites to GSH synthesis. Finally, we identify spleen tyrosine kinase (Syk) as a critical downstream signaling mediator that translates OxPL-induced effects into ceramide production and inflammatory gene regulation.ConclusionsTogether, these data demonstrate the metabolic and bioenergetic requirements that enable macrophages to translate tissue oxidation status into either antioxidant or inflammatory responses via sensing OxPL. Targeting dysregulated redox homeostasis in macrophages could therefore lead to novel therapies to treat chronic inflammation.
Oxidized phospholipids are products of lipid oxidation that are found on oxidized low-density lipoproteins and apoptotic cell membranes. These biologically active lipids were shown to affect a variety of cell types and attributed pro-as well as anti-inflammatory effects. In particular, macrophages exposed to oxidized phospholipids drastically change their gene expression pattern and function. These ‘Mox,’macrophages were identified in atherosclerotic lesions, however, it remains unclear how lipid oxidation products are sensed by macrophages and how they influence their biological function. Here, we review recent developments in the field that provide insight into the structure, recognition, and downstream signaling of oxidized phospholipids in macrophages.
Two
major subclasses of mononuclear non-heme ferrous enzymes use
two electron-donating organic cofactors (α-ketoglutarate or
pterin) to activate O2 to form FeIVO
intermediates that further react with their substrates through hydrogen
atom abstraction or electrophilic aromatic substitution. New spectroscopic
methodologies have been developed, enabling the study of the active
sites in these enzymes and their oxygen intermediates. Coupled to
electronic structure calculations, the results of these spectroscopies
provide fundamental insight into mechanism. This Perspective summarizes
the results of these studies in elucidating the mechanism of dioxygen
activation to form the FeIVO intermediate and the
geometric and electronic structure of this intermediate that enables
its high reactivity and selectivity in product formation.
Methanotrophic
bacteria utilize the nonheme diiron enzyme soluble
methane monooxygenase (sMMO) to convert methane to methanol in the
first step of their metabolic cycle under copper-limiting conditions.
The structure of the sMMO Fe(IV)2 intermediate Q responsible
for activating the inert C–H bond of methane (BDE = 104 kcal/mol)
remains controversial, with recent studies suggesting both “open”
and “closed” core geometries for its active site. In
this study, we employ nuclear resonance vibrational spectroscopy (NRVS)
to probe the geometric and electronic structure of intermediate Q
at cryogenic temperatures. These data demonstrate that Q decays rapidly
during the NRVS experiment. Combining data from several years of measurements,
we derive the NRVS vibrational features of intermediate Q as well
as its cryoreduced decay product. A library of 90 open and closed
core models of intermediate Q is generated using density functional
theory to analyze the NRVS data of Q and its cryoreduced product as
well as prior spectroscopic data on Q. Our analysis reveals that a
subset of closed core models reproduce these newly acquired NRVS data
as well as prior data. The reaction coordinate with methane is also
evaluated using both closed and open core models of Q. These studies
show that the potent reactivity of Q toward methane resides in the
“spectator oxo” of its Fe(IV)2O2 core, in contrast to nonheme mononuclear Fe(IV)O enzyme
intermediates that H atoms abstract from weaker C–H bonds.
The extradiol dioxygenases (EDOs) and intradiol dioxygenases (IDOs) are nonheme iron enzymes that catalyze the oxidative aromatic ring cleavage of catechol substrates, playing an essential role in the carbon cycle. The EDOs and IDOs utilize very different Fe II and Fe III active sites to catalyze the regiospecificity in their catechol ring cleavage products. The factors governing this difference in cleavage have remained undefined. The EDO homoprotocatechuate 2,3-dioxygenase (HPCD) and IDO protocatechuate 3,4-dioxygenase (PCD) provide an opportunity to understand this selectivity, as key O 2 intermediates have been trapped for both enzymes. Nuclear resonance vibrational spectroscopy (in conjunction with density functional theory calculations) is used to define the geometric and electronic structures of these intermediates as Fe II -alkylhydroperoxo (HPCD) and Fe III -alkylperoxo (PCD) species. Critically, in both intermediates, the initial peroxo bond orientation is directed toward extradiol product formation. Reaction coordinate calculations were thus performed to evaluate both the extra-and intradiol O−O cleavage for the simple organic alkylhydroperoxo and for the Fe II and Fe III metal catalyzed reactions. These results show the Fe II -alkylhydroperoxo (EDO) intermediate undergoes facile extradiol O−O bond homolysis due to its extra e − , while for the Fe III -alkylperoxo (IDO) intermediate the extradiol cleavage involves a large barrier and would yield the incorrect extradiol product. This prompted our evaluation of a viable mechanism to rearrange the Fe III -alkylperoxo IDO intermediate for intradiol cleavage, revealing a key role in the rebinding of the displaced Tyr447 ligand in this rearrangement, driven by the proton delivery necessary for O−O bond cleavage.
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