Ecophysiology of syntrophic communities that degrade saturated and unsaturated long-chain fatty acids

Syntrophic relationships are the key for biodegradation in methanogenic environments. We review the ecological and physiological features of syntrophic communities involved in the degradation of saturated and unsaturated long-chain fatty acids (LCFA), as well as their potential application to convert lipids/fats containing waste to biogas. Presently, about 14 species have been described with the ability to grow on fatty acids in syntrophy with methanogens, all belonging to the families Syntrophomonadaceae and Syntrophaceae . The principle pathway of LCFA degradation is through β-oxidation, but the initial steps in the conversion of unsaturated LCFA are unclear. Communities enriched on unsaturated LCFA also degrade saturated LCFA, but the opposite generally is not the case. For efficient methane formation, the physical and inhibitory effects of LCFA on methanogenesis need to be considered. LCFA adsorbs strongly to biomass, which causes encapsulation of active syntrophic communities and hampers diffusion of substrate and products in and out of the biomass. Quantification of archaea by real-time PCR analysis suggests that potential LCFA inhibitory effect towards methanogens might be reversible. Rather, the conversion of adsorbed LCFA in batch assays was shown to result in a significant increase of archaeal cell numbers in anaerobic sludge samples.     

Street heroin induces mitochondrial dysfunction and apoptosis in rat cortical neurons

Cortical function has been suggested to be highly compromised by repeated heroin self-administration. We have previously shown that street heroin induces apoptosis in neuronal-like PC12 cells. Thus, we analysed the apoptotic pathways involved in street heroin neurotoxicity using primary cultures of rat cortical neurons. Our street heroin sample was shown to be mainly composed by heroin, 6-monoacetylmorphine and morphine. Exposure of cortical neurons to street heroin induced a slight decrease in metabolic viability, without loss of neuronal integrity. Early activation of caspases involved in the mitochondrial apoptotic pathway was observed, culminating in caspase 3 activation, Poly-ADP Ribose Polymerase (PARP) cleavage and DNA fragmentation. Apoptotic morphology was completely prevented by the non-selective caspase inhibitor z-VAD-fmk, indicating an important role for caspases in neurodegeneration induced by street heroin. Ionotropic glutamate receptors, opioid receptors and oxidative stress were not involved in caspase 3 activation. Interestingly, street heroin cytotoxicity was shown to be independent of a functional mitochondrial respiratory chain, as determined using NT-2 rho(0) cells. Nonetheless, in street heroin-treated cortical neurons, cytochrome c was released, accompanied by a decrease in mitochondrial potential and Bcl-2/Bax. Pure heroin hydrochloride similarly decreased metabolic viability but only slightly activated caspase 3. Altogether, our data suggest an important role for mitochondria in mediating street heroin neurotoxic effects.     

Reducing saturated fatty acids in Arabidopsis seeds by expression of a Caenorhabditis elegans 16:0–specific desaturase

Plant oilseeds are a major source of nutritional oils. Their fatty acid composition, especially the proportion of saturated and unsaturated fatty acids, has important effects on human health. Because intake of saturated fats is correlated with the incidence of cardiovascular disease and diabetes, a goal of metabolic engineering is to develop oils low in saturated fatty acids. Palmitic acid (16:0) is the most abundant saturated fatty acid in the seeds of many oilseed crops and in Arabidopsis thaliana. We expressed FAT–5, a membrane‐bound desaturase cloned from Caenorhabditis elegans, in Arabidopsis using a strong seed‐specific promoter. The FAT‐5 enzyme is highly specific to 16:0 as substrate, converting it to 16:1?9; expression of fat‐5 reduced the 16:0 content of the seed by two‐thirds. Decreased 16:0 and elevated 16:1 levels were evident both in the storage and membrane lipids of seeds. Regiochemical analysis of phosphatidylcholine showed that 16:1 was distributed at both positions on the glycerolipid backbone, unlike 16:0, which is predominately found at the sn‐1 position. Seeds from a plant line homozygous for FAT–5 expression were comparable to wild type with respect to seed set and germination, while oil content and weight were somewhat reduced. These experiments demonstrate that targeted heterologous expression of a desaturase in oilseeds can reduce the level of saturated fatty acids in the oil, significantly improving its nutritional value.     

BMCP1: a mitochondrial uncoupling protein in neurons which regulates mitochondrial function and oxidant production

Outside the nervous system, members of the mitochondrial uncoupling protein (UCP) family have been proposed to contribute to control of body temperature and energy metabolism, and regulation of mitochondrial production of reactive oxygen species (ROS). However, the function of brain mitochondrial carrier protein 1 (BMCP1), which is highly expressed in brain, remains to be determined. To study BMCP1 expression and function in the nervous system, a high-affinity antibody to BMCP1 was generated and used to analyze tissue expression of BMCP1 protein in mouse. BMCP1 protein was highly expressed in heart and kidney, but not liver or lung. In the nervous system, BMCP1 was present in cortex, basal ganglia, substantia nigra, cerebellum, and spinal cord. Both BMCP1 mRNA and protein expression was almost exclusively neuronal. To study the effect of BMCP1 expression on mitochondrial function, neuronal (GT1-1) cell lines with stable overexpression of BMCP1 were generated. Transfected cells had higher State 4 respiration and lower mitochondrial membrane potential (psi(m)), consistent with greater mitochondrial uncoupling. BMCP1 expression also decreased mitochondrial production of ROS. These data suggest that BMCP1 can modify mitochondrial respiratory efficiency and mitochondrial oxidant production, and raise the possibility that BMCP1 might alter the vulnerability of brain to both acute injury and to neurodegenerative conditions.     

Curcumin induces concentration-dependent alterations in mitochondrial function through ROS in C2C12 mouse myoblasts

Curcumin exhibits antioxidant properties in normal cells where the uptake is low, unlike in tumor cells where uptake is high and curcumin increases reactive oxygen species (ROS) production and cell death. Mitochondria are the main source and primary target of cellular ROS. We hypothesized that curcumin would regulate cellular redox status and mitochondrial function, depending on cell sensitivity and/or curcumin concentration in normal cells. We examined the differences between low and high concentrations of curcumin, with specific attention focused on ROS levels, mitochondrial function, and cell viability in mouse C2C12 myoblast under normal and simulated conditions of diabetes. Cells incubated with high concentrations of curcumin (10–50 μM) resulted in decreased cell viability and sustained robust increases in ROS levels. Mechanistic studies showed that increased ROS levels in cells incubated with 20 μM curcumin induced opening of mitochondrial permeability transition pores and subsequent release of cytochrome c, activation of caspases 9 and 3/7, and apoptotic cell death. Low concentrations of curcumin (1–5 μM) did not affect cell viability, but induced a mild increase in ROS levels, which peaked at 2 hr after the treatment. Incubation with 5 μM curcumin also induced ROS-dependent increases in mitochondrial mass and membrane potential. Finally, pretreatment with 5 μM curcumin prevented high glucose-induced oxidative cell injury. Our study suggests that mitochondria respond differentially depending on curcumin concentration-dependent induction of ROS. The end result is either cell protection or death. Curcumin may be an effective therapeutic target for diabetes and other mitochondrial diseases when used in low concentrations.     

Mitochondrial dysfunction in diabetes and the regulatory roles of antidiabetic agents on the mitochondrial function

The prevalence of type 2 diabetes mellitus (T2DM) is increasing rapidly with its associated morbidity and mortality. Many pathophysiological pathways such as oxidative stress, inflammatory responses, adipokines, obesity-induced insulin resistance, improper insulin signaling, and beta cell apoptosis are associated with the development of T2DM. There is increasing evidence of the role of mitochondrial dysfunction in the onset of T2DM, particularly in relation to the development of diabetic complications. Here, the role of mitochondrial dysfunction in T2DM is reviewed together with its modulation by antidiabetic therapeutic agents, an effect that may be independent of their hypoglycemic effect.     

Advances in the mechanism and treatment of mitochondrial quality control involved in myocardial infarction

Mitochondria are important organelles in eukaryotic cells. Normal mitochondrial homeostasis is subject to a strict mitochondrial quality control system, including the strict regulation of mitochondrial production, fission/fusion and mitophagy. The strict and accurate modulation of the mitochondrial quality control system, comprising the mitochondrial fission/fusion, mitophagy and other processes, can ameliorate the myocardial injury of myocardial ischaemia and ischaemia-reperfusion after myocardial infarction, which plays an important role in myocardial protection after myocardial infarction. Further research into the mechanism will help identify new therapeutic targets and drugs for the treatment of myocardial infarction. This article aims to summarize the recent research regarding the mitochondrial quality control system and its molecular mechanism involved in myocardial infarction, as well as the potential therapeutic targets in the future.     

Berberine alleviates lipid metabolism disorders via inhibition of mitochondrial complex I in gut and liver

This study is to investigate the relationship between berberine (BBR) and mitochondrial complex I in lipid metabolism. BBR reversed high-fat diet-induced obesity, hepatic steatosis, hyperlipidemia and insulin resistance in mice. Fatty acid consumption, β-oxidation and lipogenesis were attenuated in liver after BBR treatment which may be through reduction in SCD1, FABP1, CD36 and CPT1A. BBR promoted fecal lipid excretion, which may result from the reduction in intestinal CD36 and SCD1. Moreover, BBR inhibited mitochondrial complex I-dependent oxygen consumption and ATP synthesis of liver and gut, but no impact on activities of complex II, III and IV. BBR ameliorated mitochondrial swelling, facilitated mitochondrial fusion, and reduced mtDNA and citrate synthase activity. BBR decreased the abundance and diversity of gut microbiome. However, no change in metabolism of recipient mice was observed after fecal microbiota transplantation from BBR treated mice. In primary hepatocytes, BBR and AMPK activator A769662 normalized oleic acid-induced lipid deposition. Although both the agents activated AMPK, BBR decreased oxygen consumption whereas A769662 increased it. Collectively, these findings indicated that BBR repressed complex I in gut and liver and consequently inhibited lipid metabolism which led to alleviation of obesity and fatty liver. This process was independent of intestinal bacteria.     

Inclusion/exclusion of fatty acids in amylose complexes as a function of the fatty acid chain length

Structural models are proposed for amylose-fatty acid complexes depending on the respective chain lengths of their constituents. The three studied fatty acids induce the Vh amylose crystalline type. However, in contrast to lauric and palmitic acids, caprylic acid is not present in crystals. On the basis of the relative amounts of amylose and fatty acid determined in complexes and previous results of molecular modelling, inclusion of lauric and palmitic acids inside the amylose helices is proposed; the acyl chains are included in crystalline areas and the car☐ylic groups in amorphous areas. The absence of caprylic acid in crystals could be due to the solubility of this compound in the crystallization medium.     

Lipid lateral diffusion and local microviscosity in plant mitochondrial membranes with various length and unsaturation of fatty acids

Two main aspects of the lipid dynamics, local microviscosity and lateral diffusion, were investigated in intact plant mitochondria isolated from different tissues exhibiting large differences in their fatty acids in terms of unsaturation (amount of linoleic and linolenic acids) or length of the hydrocarbon chains. In addition, the same parameters were determined in the outer and inner membranes isolated from cauliflower mitochondria, which differed not only in the fatty acid composition but also by the lipid-to-protein ratio. In intact mitochondria, local microviscosity assayed with anthroyloxy-fatty acids exhibited a transverse gradient from the surface to the center of the bilayer, which was mainly affected by the unsaturation index and the content in linoleic or linolenic acids. In contrast, lipid lateral diffusion increased as the content in linolenic or palmitic acids increased, but was not directly correlated to the unsaturation index. Interestingly, local microviscosity at the membrane surface was higher in the outer membrane than in the inner membrane, whereas no significant difference was found in lipid lateral diffusion. These results indicate that the influence of the fatty acid composition of mitochondrial membranes on the dynamics of the phospholipid bilayer depends on the type of movement considered and suggest that other parameters, such as the protein content of the bilayer, also affect membrane fluidity.     

Neurotoxicity of ammonia and fatty acids: Differential inhibition of mitochondrial dehydrogenases by ammonia and fatty acyl coenzyme a derivatives

In several metabolic encephalopathies, hyperammonemia and organic acidemia are consistently found. Ammonia and fatty acids (FAs) are neurotoxic: previous workers have shown that ammonia and FAs can act singly, in combination, or synergistically, in inducing coma in experimental animals. However, the biochemical mechanisms underlying the neurotoxicity of ammonia and FAs have not been fully elucidated. FAs are normally converted to their corresponding CoA derivatives (CoAs) once they enter cells and it is known that these fatty acyl CoAs can alter intermediary metabolism. The present study was initiated to determine the effects of ammonia and fatty acyl CoAs on brain mitochondrial dehydrogenases. At a pathophysiological level (2 mM), ammonia is a potent inhibitor of brain mitochondrial -ketoglutarate dehydrogenase complex (KGDHC). Only at toxicological levels (10–20 mM) does ammonia inhibit brain mitochondrial NAD⁺- and NADP⁺-linked isocitrate dehydrogenase (NAD-ICDH, NADP-ICDH), and NAD⁺-linked malate dehydrogenase (MDH) and liver mitochondrial NAD-ICDH. Butyryl- (BCoA), octanoyl- (OCoA), and palmitoyl (PCoA) CoA were potent inhibitors of brain mitochondrial KGDHC, with IC₅₀ values of 11, 20, and 25 M, respectively; moreover, the inhibitory effect of fatty acyl CoAs and ammonia were additive. At levels of 250 M or higher, both OCoA (IC₅₀=1.15 mM) and PCoA (IC₅₀=470 M) inhibit brain mitochondrial NADP-ICDH; only at higher levels (0.5–1 mM) does BCoA inhibit this enzyme (by 30–45%). Much less sensitive than KGDHC and NADP-ICDH, brain mitochondrial NAD-ICDH is only inhibited by 1 mM BCoA, OCoA, and PCoA by 22%, 35%, and 44%, respectively. Even at 1 mM, OCoA and PCoA (but not BCoA) only slightly inhibited brain mitochondrial MDH (by 23%). In the presence of toxicological levels of ammonia (20 mM) and fatty acyl CoAs (1 mM), the inhibitory effect of fatty acyl CoAs and ammonia on brain mitochondrial NAD-ICDH, NADP-ICDH, and MDH are only partially additive. These results provide some support for our hypothesis that selective inhibition of a rate-limiting and regulated enzymatic step (e.g., KGDHC) by ammonia and fatty acyl CoAs may be one of the major mechanisms underlying the neurotoxicity of ammonia and FAs. The data also suggest that the same mechanism may acocunt for the synergistic effect of ammonia and FAs in inducing coma. Since the inhibition of KGDHC by ammonia and fatty acyl CoAs occurs at pathophysiological levels, the results may assume some pathophysiological and/or pathogenetic importance in metabolic encephalopathies in which hyperammonemia and organic acidemia are persistent features.We dedicate this paper to Dr. Santiago Grisolia. Dr. Grisolia has carried out many pioneering studies in urea metabolism and ammonia toxicity. His interesting ideas have been influential in these and related fields of research. He continues to contribute significantly in unravelling the mechanisms of ammonia toxicity.     

Roles of pollination and short-chain saturated fatty acids in flower senescence

Pollination induced an immediate increase in ethylene production in Dianthus caryophyllus and Petunia hybrida. In Cymbidium, a lag of several hours was observed. In all three species, pollination induced premature flower senescence. Treatment of the stigmatic surface with aminoethoxyvinylglycine prior to pollination effectively blocked the increase in ethylene production and alleviated the detrimental effect of pollination on flower life.In all three tested species, octanoic and decanoic acids, when applied to the stigmatic surface, had no effect on ethylene production and flower life. In isolated Cymbidium lips placed with their cut base in solutions containing these fatty acids, no effects on red colouration, ethylene production, and ethylene forming enzyme activity were observed. In addition, ethylene sensitivity of isolated lips was not affected. The putative regulatory role of short-chain saturated fatty acids in (pollination-induced) flower senescence is discussed.     

Different role of saturated and unsaturated fatty acids in beta-cell apoptosis

It is believed that free fatty acids contribute to the pathogenesis of type 2 diabetes in humans. We have recently shown that lipoapoptosis of human beta-cells is specifically induced by saturated fatty acids while unsaturated had no effect. In the present study we tested the effect of co-incubation of different saturated and unsaturated free fatty acids on lipoapoptosis in beta-cells. RIN1046-38 cells and isolated human beta-cells were incubated with combinations of saturated fatty acids (palmitate, stearate) and mono- or polyunsaturated fatty acids (palmitoleate, oleate, and linoleate). Cells were incubated for 24-72 h with 1mM fatty acids. All unsaturated fatty acids tested completely prevented palmitate- or stearate-induced apoptosis of rat and human beta-cells as assessed by flow cytometric cell cycle analysis and TUNEL assay. This might suggest that apoptosis in vivo is predominantly determined by the content of unsaturated fatty acids in a mixed fatty acid pool.     

Elevated hydrostatic pressures induce apoptosis and oxidative stress through mitochondrial membrane depolarization in PC12 neuronal cells: A cell culture model of glaucoma

Abstract

Background: Despite the importance of oxidative stress and apoptosis through mitochondrial depolarization in neurodegenerative diseases, their roles in etiology of glaucoma are poorly understood. We aimed to investigate whether oxidative stress and apoptosis formation are altered in rat pheochromocytoma-derived cell line-12 (PC12) neuronal cell cultures exposed to elevated different hydrostatic pressures as a cell culture model of glaucoma. Materials: Cultured PC12 cells were subjected to 0, 15 and 70?mmHg hydrostatic pressure for 1 and 24?h. Then, the following values were analyzed: (a) cell viability; (b) lipid peroxidation and intracellular reactive oxygen species production; (c) mitochondrial membrane depolarization; (d) cell apoptosis; (e) caspase-3 and caspase-9 activities; (f) reduced glutathione (GSH) and glutathione peroxidase (GSH-Px). Results: The hydrostatic pressures (15 and 70?mmHg) increased oxidative cell damage through a decrease of GSH and GSH-Px values, and increasing mitochondrial membrane potential. Additionally, 70?mmHg hydrostatic pressure for 24?h indicated highest apoptotic effects, as demonstrated by plate reader analyses of apoptosis, caspase-3 and -9 values. Conclusion: The present data indicated oxidative stress, apoptosis and mitochondrial changes in PC12 cell line during different hydrostatic pressure as a cell culture model of glaucoma. This findings support the view that mitochondrial oxidative injury contributes early to glaucomatous optic neuropathy.     

Fenofibrate and PBA prevent fatty acid-induced loss of adiponectin receptor and pAMPK in human hepatoma cells and in hepatitis C virus-induced steatosis

Adiponectin receptors play a key role in steatosis and inflammation; however, very little is known about regulation of adiponectin receptors in liver. Here, we examined the effects of palmitate loading, endoplasmic reticulum (ER) stress, and the hypolipidemic agent fenofibrate on adiponectin receptor R2 (AdipoR2) levels and AMP-activated protein kinase (AMPK) in human hepatoma Huh7 cells and in Huh.8 cells, a model of hepatitis C-induced steatosis. Palmitate treatment reduced AdipoR2 protein and basal AMPK phosphorylation in Huh7 cells. Fenofibrate treatment preserved AdipoR2 and phosphorylated AMPK (pAMPK) levels in palmitate-treated cells accompanied by reduced triglyceride (TG) accumulation and less activation of ER stress markers CCAAT/enhancer binding (C/EBPβ) and eukaryotic translation initiation factor 2 α. ER stress agents thapsigargin and tunicamycin suppressed AdipoR2 and pAMPK levels in Huh7 cells, while fenofibrate and the chemical chaperone 4-phenylbutyrate (PBA) prevented these changes. AdipoR2 levels were lower in Huh.8 cells and fenofibrate treatment increased AdipoR2 while reducing activation of c-Jun N-terminal kinase and C/EBPβ expression without changing TG levels. Taken together, these results suggest that fatty acids and ER stress reduce AdipoR2 protein and pAMPK levels, while fenofibrate and PBA might be important therapeutic agents to correct lipid- and ER stress-mediated loss of AdipoR2 and pAMPK associated with nonalcoholic steatohepatitis.     

过度训练状态下心肌损伤线粒体机制的研究

目的：探讨过度训练状态下心肌组织损害的线粒体机制。方法：应用形态学手段和分子生化技术,观察运动后大鼠心肌组织酸性磷酸酶（ACP）、β-葡萄糖醛酸酶（β-GLU）和心肌线粒体膜通透性转换孔（肿）等指标的变化。结果：运动后心肌ACP和β-GLU活性明显增加,PTP开放增加。结论：过度训练后心肌线粒体的结构和功能发生了明显变化,这些变化可能与PTP开放增加密切相关。