Hepcidin deficiency in mice impairs white adipose tissue browning possibly due to a defect in de novo adipogenesis.

The role of iron in the two major sites of adaptive thermogenesis, namely the beige inguinal (iWAT) and brown adipose tissues (BAT) has not been fully understood yet. Body iron levels and distribution is controlled by the iron regulatory peptide hepcidin. Here, we explored iron homeostasis and thermogenic activity in brown and beige fat in wild-type and iron loaded Hepcidin KO mice. Hepcidin-deficient mice displayed iron overload in both iWAT and BAT, and preferential accumulation of ferritin in stromal cells compared to mature adipocytes. In contrast to BAT, the iWAT of Hepcidin KO animals featured with defective thermogenesis evidenced by an altered beige signature, including reduced UCP1 levels and decreased mitochondrial respiration. This thermogenic modification appeared cell autonomous and persisted after a 48 h-cold challenge, a potent trigger of thermogenesis, suggesting compromised de novo adipogenesis. Given that WAT browning occurs in both mice and humans, our results provide physiological results to interrogate the thermogenic capacity of patients with iron overload disorders.

GYY4137-Derived Hydrogen Sulfide Donates Electrons to the Mitochondrial Electron Transport Chain via Sulfide: Quinone Oxidoreductase in Endothelial Cells.

The protective effects of hydrogen sulphide (H2S) to limit oxidative injury and preserve mitochondrial function during sepsis, ischemia/reperfusion, and neurodegenerative diseases have prompted the development of soluble H2S-releasing compounds such as GYY4137. Yet, the effects of GYY4137 on the mitochondrial function of endothelial cells remain unclear, while this cell type comprises the first target cell after parenteral administration. Here, we specifically assessed whether human endothelial cells possess a functional sulfide:quinone oxidoreductase (SQOR), to oxidise GYY4137-released H2S within the mitochondria for electron donation to the electron transport chain. We demonstrate that H2S administration increases oxygen consumption by human umbilical vein endothelial cells (HUVECs), which does not occur in the SQOR-deficient cell line SH-SY5Y. GYY4137 releases H2S in HUVECs in a dose- and time-dependent fashion as quantified by oxygen consumption and confirmed by lead acetate assay, as well as AzMC fluorescence. Scavenging of intracellular H2S using zinc confirmed intracellular and intramitochondrial sulfur, which resulted in mitotoxic zinc sulfide (ZnS) precipitates. Together, GYY4137 increases intramitochondrial H2S and boosts oxygen consumption of endothelial cells, which is likely governed via the oxidation of H2S by SQOR. This mechanism in endothelial cells may be instrumental in regulating H2S levels in blood and organs but can also be exploited to quantify H2S release by soluble donors such as GYY4137 in living systems.

S100A8-mediated metabolic adaptation controls HIV-1 persistence in macrophages in vivo.

HIV-1 eradication is hindered by viral persistence in cell reservoirs, established not only in circulatory CD4+T-cells but also in tissue-resident macrophages. The nature of macrophage reservoirs and mechanisms of persistence despite combined anti-retroviral therapy (cART) remain unclear. Using genital mucosa from cART-suppressed HIV-1-infected individuals, we evaluated the implication of macrophage immunometabolic pathways in HIV-1 persistence. We demonstrate that ex vivo, macrophage tissue reservoirs contain transcriptionally active HIV-1 and viral particles accumulated in virus-containing compartments, and harbor an inflammatory IL-1R+S100A8+MMP7+M4-phenotype prone to glycolysis. Reactivation of infectious virus production and release from these reservoirs in vitro are induced by the alarmin S100A8, an endogenous factor produced by M4-macrophages and implicated in "sterile" inflammation. This process metabolically depends on glycolysis. Altogether, inflammatory M4-macrophages form a major tissue reservoir of replication-competent HIV-1, which reactivate viral production upon autocrine/paracrine S100A8-mediated glycolytic stimulation. This HIV-1 persistence pathway needs to be targeted in future HIV eradication strategies.

UCP2 silencing restrains leukemia cell proliferation through glutamine metabolic remodeling.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy derived from early T cell progenitors. Since relapsed T-ALL is associated with a poor prognosis improving initial treatment of patients is essential to avoid resistant selection of T-ALL. During initiation, development, metastasis and even in response to chemotherapy, tumor cells face strong metabolic challenges. In this study, we identify mitochondrial UnCoupling Protein 2 (UCP2) as a tricarboxylic acid (TCA) cycle metabolite transporter controlling glutamine metabolism associated with T-ALL cell proliferation. In T-ALL cell lines, we show that UCP2 expression is controlled by glutamine metabolism and is essential for their proliferation. Our data show that T-ALL cell lines differ in their substrate dependency and their energetic metabolism (glycolysis and oxidative). Thus, while UCP2 silencing decreases cell proliferation in all leukemia cells, it also alters mitochondrial respiration of T-ALL cells relying on glutamine-dependent oxidative metabolism by rewiring their cellular metabolism to glycolysis. In this context, the function of UCP2 in the metabolite export of malate enables appropriate TCA cycle to provide building blocks such as lipids for cell growth and mitochondrial respiration. Therefore, interfering with UCP2 function can be considered as an interesting strategy to decrease metabolic efficiency and proliferation rate of leukemia cells.

Use of H2O2 to Cause Oxidative Stress, the Catalase Issue.

Addition of hydrogen peroxide (H2O2) is a method commonly used to trigger cellular oxidative stress. However, the doses used (often hundreds of micromolar) are disproportionally high with regard to physiological oxygen concentration (low micromolar). In this study using polarographic measurement of oxygen concentration in cellular suspensions we show that H2O2 addition results in O2 release as expected from catalase reaction. This reaction is fast enough to, within seconds, decrease drastically H2O2 concentration and to annihilate it within a few minutes. Firstly, this is likely to explain why recording of oxidative damage requires the high concentrations found in the literature. Secondly, it illustrates the potency of intracellular antioxidant (H2O2) defense. Thirdly, it complicates the interpretation of experiments as subsequent observations might result from high/transient H2O2 exposure and/or from the diverse possible consequences of the O2 release.

Hypertonic external medium represses cellular respiration and promotes Warburg/Crabtree effect.

Hyperosmotic conditions are associated to several pathological states. In this article, we evaluate the consequence of hyperosmotic medium on cellular energy metabolism. We demonstrate that exposure of cells to hyperosmotic conditions immediately reduces the mitochondrial oxidative phosphorylation rate. This causes an increase in glycolysis, which represses further respiration. This is known as the Warburg or Crabtree effect. In addition to aerobic glycolysis, we observed two other cellular responses that would help to preserve cellular ATP level and viability: A reduction in the cellular ATP turnover rate and a partial mitochondrial uncoupling which is expected to enhance ATP production by Krebs cycle. The latter is likely to constitute another metabolic adaptation to compensate for deficient oxidative phosphorylation that, importantly, is not dependent on glucose.

Positive feedback during sulfide oxidation fine-tunes cellular affinity for oxygen.

UNLABELLED: Sulfide (H2S in the gas form) is the third gaseous transmitter found in mammals. However, in contrast to nitric oxide (NO) or carbon monoxide (CO), sulfide is oxidized by a sulfide quinone reductase and generates electrons that enter the mitochondrial respiratory chain arriving ultimately at cytochrome oxidase, where they combine with oxygen to generate water. In addition, sulfide is also a strong inhibitor of cytochrome oxidase, similar to NO, CO and cyanide. The balance between the electron donor and the inhibitory role of sulfide is likely controlled by sulfide and oxygen availability. The present study aimed to evaluate if and how sulfide release and oxidation impacts on the cellular affinity for oxygen. RESULTS: i) when sulfide delivery approaches the maximal sulfide oxidation rate cells become exquisitely dependent on oxygen; ii) a positive feedback makes the balance between sulfide-releasing and -oxidizing rates the relevant parameter rather than the absolute values of these rates, and; iii) this altered dependence on oxygen is detected with sulfide concentrations that remain in the low micromolar range. CONCLUSIONS: i) within the context of continuous release of sulfide stemming from cellular metabolism, alterations in the activity of the sulfide oxidation pathway fine-tunes the cell's affinity for oxygen, and; ii) a decrease in the expression of the sulfide oxidation pathway greatly enhances the cell's dependence on oxygen concentration.

Mitochondrial retrograde signaling mediated by UCP2 inhibits cancer cell proliferation and tumorigenesis.

Cancer cells tilt their energy production away from oxidative phosphorylation (OXPHOS) toward glycolysis during malignant progression, even when aerobic metabolism is available. Reversing this phenomenon, known as the Warburg effect, may offer a generalized anticancer strategy. In this study, we show that overexpression of the mitochondrial membrane transport protein UCP2 in cancer cells is sufficient to restore a balance toward oxidative phosphorylation and to repress malignant phenotypes. Altered expression of glycolytic and oxidative enzymes mediated the effects of this metabolic shift. Notably, UCP2 overexpression increased signaling from the master energy-regulating kinase, adenosine monophosphate-activated protein kinase, while downregulating expression of hypoxia-induced factor. In support of recent new evidence about UCP2 function, we found that UCP2 did not function in this setting as a membrane potential uncoupling protein, but instead acted to control routing of mitochondria substrates. Taken together, our results define a strategy to reorient mitochondrial function in cancer cells toward OXPHOS that restricts their malignant phenotype.

The level of H2O2 type oxidative stress regulates virulence of Theileria-transformed leukocytes.

Theileria annulata infects predominantly macrophages, and to a lesser extent B cells, and causes a widespread disease of cattle called tropical theileriosis. Disease-causing infected macrophages are aggressively invasive, but this virulence trait can be attenuated by long-term culture. Attenuated macrophages are used as live vaccines against tropical theileriosis and via their characterization one gains insights into what host cell trait is altered concomitant with loss of virulence. We established that sporozoite infection of monocytes rapidly induces hif1-α transcription and that constitutive induction of HIF-1α in transformed leukocytes is parasite-dependent. In both infected macrophages and B cells induction of HIF-1α activates transcription of its target genes that drive host cells to perform Warburg-like glycolysis. We propose that Theileria-infected leukocytes maintain a HIF-1α-driven transcriptional programme typical of Warburg glycolysis in order to reduce as much as possible host cell H2 O2 type oxidative stress. However, in attenuated macrophages H2O2 production increases and HIF-1α levels consequently remained high, even though adhesion and aggressive invasiveness diminished. This indicates that Theileria infection generates a host leukocytes hypoxic response that if not properly controlled leads to loss of virulence.

Regulation of mitochondrial bioenergetic function by hydrogen sulfide. Part I. Biochemical and physiological mechanisms.

Until recently, hydrogen sulfide (H2 S) was exclusively viewed a toxic gas and an environmental hazard, with its toxicity primarily attributed to the inhibition of mitochondrial Complex IV, resulting in a shutdown of mitochondrial electron transport and cellular ATP generation. Work over the last decade established multiple biological regulatory roles of H2 S, as an endogenous gaseous transmitter. H2 S is produced by cystathionine γ-lyase (CSE), cystathionine ß-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3-MST). In striking contrast to its inhibitory effect on Complex IV, recent studies showed that at lower concentrations, H2 S serves as a stimulator of electron transport in mammalian cells, by acting as a mitochondrial electron donor. Endogenous H2 S, produced by mitochondrially localized 3-MST, supports basal, physiological cellular bioenergetic functions; the activity of this metabolic support declines with physiological aging. In specialized conditions (calcium overload in vascular smooth muscle, colon cancer cells), CSE and CBS can also associate with the mitochondria; H2 S produced by these enzymes, serves as an endogenous stimulator of cellular bioenergetics. The current article overviews the biochemical mechanisms underlying the stimulatory and inhibitory effects of H2 S on mitochondrial function and cellular bioenergetics and discusses the implication of these processes for normal cellular physiology. The relevance of H2 S biology is also discussed in the context of colonic epithelial cell physiology: colonocytes are exposed to high levels of sulfide produced by enteric bacteria, and serve as a metabolic barrier to limit their entry into the mammalian host, while, at the same time, utilizing it as a metabolic 'fuel'.

New biologically active hydrogen sulfide donors.

Generous donors: The dithioperoxyanhydrides (CH3 COS)2 , (PhCOS)2 , CH3 COSSCO2 Me and PhCOSSCO2 Me act as thiol-activated hydrogen sulfide donors in aqueous buffer solution. The most efficient donor (CH3 COS)2 can induce a biological response in cells, and advantageously replace hydrogen sulfide in ex vivo vascular studies.