Targeting microbiota-mitochondria inter-talk: Microbiota control mitochondria metabolism.

Our aim is to highlight the subtle relationship that exists between microbiota and mitochondria. Microbiota targets mitochondria by modulating the Reactive Oxygen Species (ROS) production and the mitochondrial activity through interactions with toxins, proteins or other metabolites released by gut microbiota. The intriguing relationship that exists between mitochondria and microbiota is strengthened by the probable prokaryotic origin of mitochondria. Emerging data implicates a role for ROS, nitric oxide, Short Chain Fatty Acids and hydrogen sulfide in the cross-talk between microbiota - mitochondria and REDOX signaling. Several studies have shown that microbiota act and modulate mitochondrial activity, and use it as a relay to strengthen host-microbiotal interaction. This modulation depends on the gut bacterial strain quality and diversity to increase its pathogenic versus beneficial effects. Furthermore, based on conclusions from new studies, it is possible that microbiota can directly interact with the host cell gene expression by favoring bacterial and mitochondrial DNA insertion in the nuclear genome. The emerging knowledge of mitochondria-microbiota interaction may be of great importance to better understand the mechanism of mitochondrial and metabolic diseases, and the syndromes associated with change in quality and quantity of microbiotal species. We suggest that microbiota via mitochondrial modulation influence cell homeostasis and metabolism. The challenge will be to find strategies to modulate the quality and diversity of microbiota rather than acting on microbiota metabolites and microbiota related factors. The medicine of tomorrow will be completely personalized. Firstly there will be a test to show the quality, quantity and diversity of microbiota, and secondly a preventive or therapeutic strategy will be administrated (probiotics, diet, prodrug or fecal transplantation). The era of digital medicine is here.

Maillard reaction, mitochondria and oxidative stress: potential role of antioxidants.

Glycation and oxidative stress are two important processes known to play a key role in complications of many disease processes. Oxidative stress, either via increasing reactive oxygen species (ROS), or by depleting the antioxidants may modulate the genesis of early glycated proteins in vivo. Maillard Reactions, occur in vivo as well as in vitro and are associated with the chronic complications of diabetes, aging and age-related diseases. Hyperglycaemia causes the autoxidation of glucose, glycation of proteins, and the activation of polyol metabolism. These changes facilitate the generation of reactive oxygen species and decrease the activity of antioxidant enzymes such as Cu,Zn-superoxide dismutase, resulting in a remarkable increase of oxidative stress. A large body of evidence indicates that mitochondria alteration is involved and plays a central role in various oxidative stress-related diseases. The damaged mitochondria produce more ROS (increase oxidative stress) and less ATP (cellular energy) than normal mitochondria. As they are damaged, they cannot burn or use glucose or lipid and cannot provide cell with ATP. Further, glucose, amino acids and lipid will not be correctly used and will accumulate outside the mitochondria; they will undergo more glycation (as observed in diabetes, obesity, HIV infection and lipodystrophia). The objective of this paper is to discuss how to stop the vicious circle established between oxidative stress, Maillard Reaction and mitochondria. The potential application of some antioxidants to reduce glycation phenomenon and to increase the antioxidant defence system by targeting mitochondria will be discussed. Food and pharmaceutical companies share the same challenge, they must act now, urgently and energetically.

Neurodegenerative diseases and oxidative stress.

Oxidative stress is now recognized as accountable for redox regulation involving reactive oxygen species (ROS) and reactive nitrogen species (RNS). Its role is pivotal for the modulation of critical cellular functions, notably for neurons astrocytes and microglia, such as apoptosis program activation, and ion transport, calcium mobilization, involved in excitotoxicity. Excitotoxicity and apoptosis are the two main causes of neuronal death. The role of mitochondria in apoptosis is crucial. Multiple apoptotic pathways emanate from the mitochondria. The respiratory chain of mitochondria that by oxidative phosphorylation, is the fount of cellular energy, i.e. ATP synthesis, is responsible for most of ROS and notably the first produced, superoxide anion (O(2)(;-)). Mitochondrial dysfunction, i.e. cell energy impairment, apoptosis and overproduction of ROS, is a final common pathogenic mechanism in aging and in neurodegenerative disease such as Alzheimer's disease (AD), Parkinson's disease (PD) and amyotrophic lateral sclerosis (ALS). Nitric oxide (NO(;)), an RNS, which can be produced by three isoforms of NO-synthase in brain, plays a prominent role. The research on the genetics of inherited forms notably ALS, AD, PD, has improved our understanding of the pathobiology of the sporadic forms of neurodegenerative diseases or of aging of the brain. ROS and RNS, i.e. oxidative stress, are not the origin of neuronal death. The cascade of events that leads to neurons, death is complex. In addition to mitochondrial dysfunction (apoptosis), excitotoxicity, oxidative stress (inflammation), the mechanisms from gene to disease involve also protein misfolding leading to aggregates and proteasome dysfunction on ubiquinited material.

Could antioxidant supplementation reduce antiretroviral therapy-induced chronic stable hyperlactatemia?

OBJECTIVE: To determine if asymptomatic stable chronic hyperlactatemia in human immunodeficiency virus (HIV)-infected patients under highly active antiretroviral therapy (HAART, including nucleoside analog reverse transcriptase inhibitors (NRTI)) could be improved by antioxidant supplementation. DESIGN: To match two groups of patients taking NRTI for at least 24 months: 15 without and 15 with antioxidant supplementation (vitamin E, beta-carotene, N-acetylcysteine, selenium, Gingko biloba extracts and nutritional supplements). For both the groups, the supplementation by antioxidants or its lack was carefully assessed. Venous lactatemia, blood oxidative stress markers (plasma lipid peroxidation, enzymatic and non-enzymatic antioxidants), CDC revisited classification, CD4 count and viral load, NRTI (with or without stavudine) and other antiretroviral drugs used, lipoatrophy, central fat accumulation were assessed. RESULTS: Patients were not statistically different with respect to the CDC classification, CD4 count, viral load and characteristics of antiretroviral therapy. Blood oxidative stress markers, i.e. vitamin E, vitamin A and beta-carotene tended to be higher in the supplemented group. The difference observed in venous lactate concentration between the two groups was significant (1.37 +/- 0.10 vs. 1.82 +/- 0.19 mmol/l in the supplemented and non-supplemented groups, respectively P = 0.04). CONCLUSION: Antioxidant supplementation improves the asymptomatic stable chronic hyperlactatemia observed in HIV-infected patients taking HAART including NRTI for a long time. Controlled studies are needed to demonstrate the efficacy of this supplementation on mitochondrial toxicity observed during HAART and the possible usefulness of its combination with mitochondrial cofactors like carnitine, riboflavine, coenzyme Q, alpha-lipoic acid.

Effect of ethyl esterification of phenolic acids on low-density lipoprotein oxidation.

Inhibition of copper-induced low-density lipoprotein (LDL) oxidation by phenolic acids and their ethyl esters was investigated. LDL oxidation was evaluated by the hydroperoxide concentration and the chromatographic pattern of apoprotein fractions after fast protein liquid chromatography (FPLC). Antiradical properties against 1,1-diphenyl-2-picryl hydrazyl (DPPH) radical and 2,2'-azobis(2-amidinopropane)dihydrochloride (AAPH) were also investigated, and lipophilicity determined by thin-layer chromatography. Caffeic acid at 5 microM and sinapic acid at 10 microM protected LDL against oxidation, inhibiting both hydroperoxide formation and the increase of apoprotein negative charge. Ferulic, gallic and p-hydroxy cinnamic acids were ineffective. Ethyl esterification increased the lipophilicity of the five acids, and enhanced the antioxidant properties of caffeic, sinapic and ferulic acids. Ethyl caffeate was protective at 1 microM. In contrast, gallic and p-hydroxy cinnamic ethyl esters were ineffective. Our results indicate that ethyl esterification of phenolic acids increases lipophilicity of their ethyl esters and may enable a better incorporation into the lipid layer of the LDL particle and the exertion of their antioxidant effect in the true site of lipoperoxidation. However, increasing lipophilicity is not the only mechanism able to potentiate preexisting antioxidant properties of molecules, and probably other mechanisms are implicated.

Oxidized-LDL induce apoptosis in HUVEC but not in the endothelial cell line EA.hy 926.

We studied the cytotoxic effect of copper-oxidized LDL in human primary human umbilical vein endothelial cells (HUVEC) and the immortalized EA.hy 926 cell line. Copper oxidized LDL (50-200 microg apoB/ml) induced concentration-dependent apoptotic cell death in HUVEC but did not induce apoptosis in EA.hy 926 cells. Only necrotic EA.hy 926 cells were evidenced at all copper oxidized LDL concentrations (25-200 microg apoB/ml), oxidation states (lightly, moderately and extensively copper-oxidized LDL) and incubation periods (4, 8 and 20 h). The different mechanisms of cell death induced by copper-oxidized LDL in EA.hy 926 cells and HUVEC may be related to various factors such as cytokines. In this study, we investigated whether interleukin-8 may be implicated in this process. The interleukin-8 production was increased in EA.hy 926 cells but not in HUVEC incubated with oxidized LDL. This increase in EA.hy 926 cells was associated with necrosis but not apoptosis. Nevertheless, the addition of interleukin-8 to HUVEC did not inhibit apoptosis induced by oxidized LDL. As the lower antioxidant capacity of EA.hy 926 cells results in higher sensitivity to oxidized LDL cytotoxicity (as we previously described), the redox status of cells may also control the form of endothelial cell death. In atherosclerotic lesions, the formation of apoptotic endothelial cells may result in part from the induction by oxidized LDL.

Protective effects of the lipophilic redox conjugate tocopheryl succinyl-ethyl ferulate on HIV replication.

Previously, we demonstrated that ferulate ethyl and tocopherol reduced HIV replication. In this study, we investigate whether the conjugation of both compounds (O-tocopheryl succinyl O-ethyl ferulate) can increase HIV inhibition. We show here for the first time that O-tocopheryl succinyl O-ethyl ferulate inhibits 80% of HIV replication (HIV-1 acute infection and HIV transmission), inhibits cell lipoperoxidation and prevents cellular glutathione consumption. Compared to ferulate ethyl and tocopheryl succinyl, O-tocopheryl succinyl O-ethyl ferulate inhibits more HIV replication. This may be due in part to the great increase in the lipophilicity of this compound.

Comparison of oxidized low-density lipoprotein toxicity on EA.hy 926 cells and human vein endothelial cells: influence of antioxidant systems.

We compared the susceptibility to oxidized LDL cytotoxicity of primary human umbilical vein endothelial cells (HUVEC) and EA.hy 926 cells. EA.hy 926 endothelial cells were more susceptible than HUVEC. To determine the basis of this difference, we evaluated the enzymatic antioxidant machinery in the two cell types. The antioxidant enzyme activities of superoxide dismutase, catalase and glutathione peroxidase were significantly lower in EA.hy cells than in HUVEC: 54%, 71% and 8% of the HUVEC enzyme activities respectively. Pre-incubation of the EA.hy 926 endothelial cells with glutathione peroxidase (100 IU/ml) inhibited the cytotoxic effect of oxidized LDL. Superoxide dismutase (300 or 600 IU/ml) and catalase (300 or 600 IU/ml) had no effect. Compared to HUVEC, the higher susceptibility of EA.hy 926 cells to oxidized LDL induced injury may be associated with lower antioxidant defences, in particular with lower glutathione peroxidase activity which is known to eliminate lipid hydroperoxides and thereby to prevent the formation of damaging peroxyl radical intermediates.

Clastogenic factors in plasma of HIV-1 infected patients activate HIV-1 replication in vitro: inhibition by superoxide dismutase.

The frequent neoplastic disorders present in HIV-infected patients and the implication of oxidative stress in AIDS-Kaposi's sarcoma pathogenesis prompted us to study whether the mechanisms implicated in genotoxic effects of clastogenic factors (CFs) (i.e., chromosome damaging materials released by cells under conditions of oxidant stress) can play a role in HIV-1 expression and whether exogenous superoxide dismutase can inhibit the clastogenic and HIV-inducing effects of CFs. CFs were found in the plasma of all HIV-1 infected patients (n = 21) of this study group, in asymptomatic (CDC II) as well as in symptomatic patients (CDC IV). In addition to their chromosome damaging effect, CFs are able to upregulate HIV-1 expression in U1 cells and in PBMCs activated with PHA and IL2 at all time points (p < .05). Their formation, therefore, is an early event in the disease. It occured despite antiviral medication in these patients. Superoxide dismutase inhibited the clastogenic and the viral inducing effects (p < .05). On the basis of our findings, association of SOD mimetics or superoxide scavengers with antiviral drugs may be a new therapeutic approach. This polytherapy, if started early enough after infection, may prolong the latency period and limit the emergence of drug-resistant viral strains.

Phenylarsine oxide inhibits ex vivo HIV-1 expression.

Phenylarsine oxide (PAO), which is described as an inhibitor of tyrosine phosphatase activity, inhibits H2O2 release from human peripheral blood mononuclear cells (PBMCs) as measured by electrochemistry. Since human immunodeficiency virus type 1 (HIV-1) replication is known to be favored under oxidative stress conditions, ex vivo experiments using uninfected PBMCs, primary monocytes or a latently infected promonocytic U1 cell line show that HIV-1 replication and reactivation, monitored by p24 antigen measurement, are inhibited by PAO in a time- and concentration-dependent manner. These observations can be linked with the inhibition of NF-kappa B activation when uninfected monocytes are induced by either tumor necrosis factor alpha (TNF-alpha) phorbol 12-myristate 13-acetate (PMA) or lipopolysaccharide (LPS).

Immunocytochemical study of uptake of exogenous carrier-free copper-zinc superoxide dismutase by peripheral blood lymphocytes.

We recently demonstrated that exogenous copper-zinc superoxide dismutase (SOD) reduced HIV replication in tumor necrosis factor alpha activated chronically HIV-infected promonocytic U1 cell line and in peripheral blood mononuclear cells coculture. However, whether exogenous SOD penetrates the cellular membrane or acts extracellularly has been remained controversial. SOD has been considered as not to penetrate the cellular membrane because of its high molecular weight, thus the main site of action is presumed to be extracellular. In order to determine whether exogenous SOD penetrates inside the cell, we utilized a gentle immunocytochemical method to detect Mn and Cu,Zn SOD in peripheral blood lymphocytes incubated with various concentrations of exogenous carrier-free Cu,Zn SOD without prior permeabilization of cell membranes. After 24 hrs. the total SOD activity and immunocytochemical studies were performed. Here we demonstrate clearly that a large amount of carrier-free Cu,Zn SOD, added exogenously, penetrates the cellular membrane and increases total SOD activity.

Oxidized low-density lipoprotein induces the production of interleukin-8 by endothelial cells.

The concentration of interleukin-8 (IL-8) and RANTES was measured in culture supernatants of human EA.hy 926 endothelial cells incubated with oxidized low-density lipoproteins (LDL). Oxidized LDL induced a 3-fold increase in IL-8 production (p < 0.01), whereas RANTES was not detected. Native LDL did not stimulate IL-8 production. IL-8 production in oxidized-LDL-treated cells was mediated by reactive oxygen species, as it was partially inhibited by catalase and completely inhibited by glutathione peroxidase and N-acetylcysteine (p < 0.01).

[Effect of the liposolubility of free radical scavengers on the production of antigen P24 from a HIV infected monocytic cell line]. / Influence de la liposolubilité des molécules antiradicalaires sur la production d'antigène P24 par des cellules monocytaires infectées par le VIH.

A large body of evidence indicates that AIDS may be the consequence of a virus-induced antioxidant deficiency and implicates reactive oxygen species (ROS) in the pathogenesis of HIV infection. The high level of antigenic acid and cytokines activities in AIDS results in the production of superoxides (O2-), hydrogen peroxide (H202) and hydroxyl radicals (OH). HIV-infected T cells display low levels of SOD, catalase, thioredoxin and glutatione peroxidase rendering them susceptible to undergo apoptosis. Induction of NF kappa B and HIV replication are at least in part dependent on reactive oxygen intermediates. We examined the protective effects of two antioxidants. Ferulic Acid (FA) and Ethyl Ferulate (EF) at 1, 5, 10, 10 microM on the TNF induced HIV activation in the chronically infected promonocytic U1 cell line. FA and EF at 5 microM elicit a marked decrease of HIV p24 release. HIV inhibition was greater after pretreatment with EF than with FA. At these concentrations, no cytotoxicity was observed. When SOD (100 UI) was combined with EF and FA no more inhibition was observed. But when SOD was added alone, it induced a marked inhibition (30%). This class of drugs may present potential interest as antiviral agents or as adjuvant therapy in AIDS.

Inhibition of lung epithelial cell proliferation by hyperoxia. Posttranscriptional regulation of proliferation-related genes.

The alveolar surface of the lung is a major target for oxidant injury. After injury, repair of the alveolar epithelium is dependent on the ability of epithelial type 2 (T2) cells to proliferate. The regulation of T2 cell proliferation and the effect of reactive oxygen (O2) species on this lung cell proliferation have not been well defined. To investigate this process we focused on the regulation of two late cell cycle genes, histone and thymidine kinase, in T2 cells and fibroblasts exposed in vitro to varying periods of hyperoxia (95% O2). Hyperoxia for 24 to 48 h arrested cell proliferation in a SV40T-immortalized T2 cell line we have developed and in primary and SV40T-immortalized lung fibroblasts. Despite the cessation of proliferation, histone and TK mRNA continued to be expressed at high levels; mRNA half-lives were markedly prolonged but neither protein was translated. Thus proliferation arrest induced by hyperoxia was associated with posttranscriptional control of at least two late cell cycle-related genes. This form of proliferation arrest is also seen when primary and SV40T-T2 cells but not fibroblasts are serum deprived, suggesting that T2 cells in vitro may be uniquely sensitive to alterations in their redox state and that these alterations in turn affect translational control of a subset of proliferation-related genes.