Linkage of Oxidative Stress and Mitochondrial Dysfunctions to Spontaneous Culture Degeneration in Aspergillus nidulans

Filamentous fungi including mushrooms frequently and spontaneously degenerate during subsequent culture maintenance on artificial media, which shows the loss or reduction abilities of asexual sporulation, sexuality, fruiting, and production of secondary metabolites, thus leading to economic losses during mass production. To better understand the underlying mechanisms of fungal degeneration, the model fungus Aspergillus nidulans was employed in this study for comprehensive analyses. First, linkage of oxidative stress to culture degeneration was evident in A. nidulans. Taken together with the verifications of cell biology and biochemical data, a comparative mitochondrial proteome analysis revealed that, unlike the healthy wild type, a spontaneous fluffy sector culture of A. nidulans demonstrated the characteristics of mitochondrial dysfunctions. Relative to the wild type, the features of cytochrome c release, calcium overload and up-regulation of apoptosis inducing factors evident in sector mitochondria suggested a linkage of fungal degeneration to cell apoptosis. However, the sector culture could still be maintained for generations without the signs of growth arrest. Up-regulation of the heat shock protein chaperones, anti-apoptotic factors and DNA repair proteins in the sector could account for the compromise in cell death. The results of this study not only shed new lights on the mechanisms of spontaneous degeneration of fungal cultures but will also provide alternative biomarkers to monitor fungal culture degeneration.Culture degeneration, also called colony deterioration, of filamentous fungi can frequently occur during subsequent maintenance of fungal culture on artificial media by showing morphological changes, such as colony sectorization, loss or impaired ability of sporulation, fruiting, and sexuality (1, 2). It was first called as “woolly degeneration” in the model fungus Neurospora crassa (3). These morphological variations are also accompanied with the loss or reduction in secondary metabolites production, thus resulting in great commercial losses (4–7). Different from the mutation of genes involved in conidiation producing fluffy phenotypes (8–10), fungal culture degeneration spontaneously occurs and is usually irreversible. The rate of colony deterioration varies considerably among fungal species/strains and correlates with the growth environments, especially the composition of nutrient medium (11, 12). In addition, unlike the phenotype of senescence observed in the model fungus Podospora anserina (13), morphological variants of other filamentous fungi could be subcultured for many generations without growth arrest (14, 15). The mechanism(s) underlying fungal culture degeneration is poorly understood. Genetic mutations were not evident in the degenerated fungal isolates of Penicillium chrysogenum (6). Otherwise, methylation of genomic DNA was reported in a sector of the plant pathogenic fungus Fusarium oxysporum after successive subculturing (16). In some fungi, degeneration was linked to chromosome instability (4, 17).Our previous studies on the insect pathogenic fungus Metarhizium anisopliae revealed that fungal culture degeneration showed the signs of aging such as cellular accumulation of reactive oxygen species (ROS)¹, mitochondrial (mt) dysfunctions, and mtDNA glycation (14, 15). According to the vicious cycle theory of aging, mitochondria are the primary source of intracellular ROS production and also one of the important targets of ROS damage, which leads to generation of additional ROS (18, 19). Mitochondrial proteomics analyses have been frequently performed for studies on human diseases, and the physiologies of plants and yeasts (20, 21). Proteins localized in the mitochondria control mt dynamics, morphology, and function and their dysregulation or damage may induce abnormality in mitochondrial function (22). However, it is not known whether these changes occur in degenerated fungal cultures.In this study, cell biology, biochemical and comparative mitochondrial proteomic analyses were performed by using the model fungus Aspergillus nidulans to better understand the features and mechanisms of fungal culture degeneration. We found a significant difference in mitochondrial protein profiles between the wild type (WT) and nonsporulation sector culture. Many of the altered proteins fall into the functional categories of energy metabolism, stress responses and cell death. Functional consequences of these changes are supported by our experimental data. The observed features such as cellular oxidative stress, mitochondrial dysfunctions, accelerated autophagy and releases of apoptotic factors in degenerated A. nidulans resemble the apoptotic process observed in mammalian cells.     

Prenatal Stress Causes Oxidative Damage to Mitochondrial DNA in Hippocampus of Offspring Rats

Mitochondrion, the primary source of reactive oxygen species (ROS), is also the target of ROS. 8-Hydroxy-2′-deoxyguanosine (8-OH-dG) is the major end-product of damaged DNA caused by ROS. In our previous studies, we showed that prenatal stress (PNS) preferentially caused cognitive dysfunction and increased ROS in the hippocampus of female offspring rats. The present study aimed to determine 8-OH-dG level of mitochondria in order to elucidate the mechanism of hippocampal pyramidal neuronal damage and cognitive dysfunction induced by PNS. Pregnant rats were divided into two groups: control group (undisturbed) and PNS group (exposed to a restraint stress for 7 days at the late stage of gestation). Offspring rats were divided into four groups: female-control group, male-control group, female-stress group, male-stress group and used at 30-day-old after their birth. The content of 8-OH-dG was determined by high performance liquid chromatography-electrochemical detection (HPLC-ECD). The results showed that the contents of 8-OH-dG in female and male prenatal stressed offspring were significantly higher than that in their respective controls (P < 0.001). 8-OH-dG level was significantly higher in the female-stress group than in the male-stress group (P < 0.05), whereas there was no any gender-dependent difference in the control groups. These results suggest that accumulation of oxidative mitochondrial DNA damage may play an important role in PNS-induced cognitive dysfunction in female offspring rats. Special issue article in honor of Dr. Akitane Mori.     

线粒体钙单向转运体和线粒体应激关系的研究进展

钙离子(calcium ion,Ca~(2+))在线粒体功能障碍及细胞损伤凋亡过程中发挥重要的细胞信号作用。近些年来关于Ca~(2+)通道以及其调控蛋白的研究越来越多,其中,线粒体单向转运体(uniporter)复合物的结构组成及其相关蛋白的分布特点成为主要研究热点。作为uniporter复合物中关键的通道蛋白,线粒体钙单向转运蛋白(mitochondrial calcium uniporter,MCU)可顺电化学梯度摄入Ca~(2+),将Ca~(2+)从胞质转运到线粒体基质并控制转运速率,其在胞内Ca~(2+)信号转导、Ca~(2+)稳态、线粒体能量代谢以及细胞凋亡方面具有重要意义。识别调控线粒体内Ca~(2+)信号的MCU及其相关蛋白可深入阐明线粒体应激在相关疾病中的发生发展,并为进一步的疾病治疗提供理论依据。     

Ablation of Ceramide Synthase 2 Causes Chronic Oxidative Stress Due to Disruption of the Mitochondrial Respiratory Chain

Ceramide is a key intermediate in the pathway of sphingolipid biosynthesis and is an important intracellular messenger. We recently generated a ceramide synthase 2 (CerS2) null mouse that cannot synthesize very long acyl chain (C22-C24) ceramides. This mouse displays severe and progressive hepatopathy. Significant changes were observed in the sphingolipid profile of CerS2 null mouse liver, including elevated C16-ceramide and sphinganine levels in liver and in isolated mitochondrial fractions. Because ceramide may be involved in reactive oxygen species (ROS) formation, we examined whether ROS generation was affected in CerS2 null mice. Levels of a number of anti-oxidant enzymes were elevated, as were lipid peroxidation, protein nitrosylation, and ROS. ROS were generated from mitochondria due to impaired complex IV activity. C16-ceramide, sphingosine, and sphinganine directly inhibited complex IV activity in isolated mitochondria and in mitoplasts, whereas other ceramide species, sphingomyelin, and diacylglycerol were without effect. A fluorescent analog of sphinganine accumulated in mitochondria. Heart mitochondria did not display a substantial alteration in the sphingolipid profile or in complex IV activity. We suggest that C16-ceramide and/or sphinganine induce ROS formation through the modulation of mitochondrial complex IV activity, resulting in chronic oxidative stress. These results are of relevance for understanding modulation of ROS signaling by sphingolipids.     

Oxidative Stress,Mitochondrial Dysfunction,and Epilepsy

Epilepsy is a common and heterogeneous neurological disorder arising from biochemical and molecular events that are incompletely understood. To effectively manage epilepsies, it is important to understand the mechanisms underlying both seizure-induced brain damage as well as seizure initiation. Oxidative stress is emerging as a mechanism that may play an important role in the etiology of seizure-induced neuronal death. Conversely, epileptic seizures are a common occurrence in mitochondrial diseases arising from defects in oxidative phosphorylation. This review focuses on the emerging role of oxidative stress and mitochondrial dysfunction both as a consequence and cause of epileptic seizures.     

Mycotoxin-Containing Diet Causes Oxidative Stress in the Mouse

Mycotoxins which mainly consist of Aflatoxin (AF), Zearalenone (ZEN) and Deoxynivalenol (DON) are commonly found in many food commodities. Although each component has been shown to cause liver toxicity and oxidative stress in several species, there is no evidence regarding the effect of naturally contained multiple mycotoxins on tissue toxicity and oxidative stress in vivo. In the present study, mycotoxins-contaminated maize (AF 597 µg/kg, ZEN 729 µg/kg, DON 3.1 mg/kg maize) was incorporated into the diet at three different doses (0, 5 and 20%) to feed the mice, and blood and tissue samples were collected to examine the oxidative stress related indexes. The results showed that the indexes of liver, kidney and spleen were all increased and the liver and kidney morphologies changed in the mycotoxin-treated mice. Also, the treatment resulted in the elevated glutathione peroxidase (GPx) activity and malondialdehyde (MDA) level in the serum and liver, indicating the presence of the oxidative stress. Moreover, the decrease of catalase (CAT) activity in the serum, liver and kidney as well as superoxide dismutase (SOD) activity in the liver and kidney tissue further confirmed the occurrence of oxidative stress. In conclusion, our data indicate that the naturally contained mycotoxins are toxic in vivo and able to induce the oxidant stress in the mouse.     

Two Light-Induced Processes in the Photoreceptor Cells of Limulus Ventral Eye

The dark-adapted current-voltage (I-V) curve of a ventral photoreceptor cell of Limulus, measured by a voltage-clamp technique, has a high slope-resistance region more negative than resting voltage, a lower slope-resistance region between resting voltage and zero, and a negative slope-resistance region more positive than 0 v. With illumination, we find no unique voltage at which there is no light-induced current. At the termination of illumination, the I-V curve changes quickly, then recovers very slowly to a dark-adapted configuration. The voltage-clamp currents during and after illumination can be interpreted to arise from two separate processes. One process (fast) changes quickly with change in illumination, has a reversal potential at +20 mv, and has an I-V curve with positive slope resistance at all voltages. These properties are consistent with a light-induced change in membrane conductance to sodium ions. The other process (slow) changes slowly with changes in illumination, generates light-activated current at +20 mv, and has an I-V curve with a large region of negative slope resistance. The mechanism of this process cannot as yet be identified.     

Neuroprotectin D1 is Synthesized in the Cone Photoreceptor Cell Line 661W and Elicits Protection Against Light-Induced Stress

Docosahexaenoic acid (DHA), an omega-3 fatty acid family member, is obtained by diet or synthesized from dietary essential omega-3 linolenic acid and delivered systemically to the choriocapillaris, from where it is taken up by the retinal pigment epithelium (RPE). DHA is then transported to the inner segments of photoreceptors, where it is incorporated in phospholipids during the biogenesis of outer segment disk and plasma membranes. As apical photoreceptor disks are gradually shed and phagocytized by the RPE, DHA is retrieved and recycled back to photoreceptor inner segments for reassembly into new disks. Under uncompensated oxidative stress, the docosanoid neuroprotectin D1 (NPD1), a potent mediator derived from DHA, is formed by the RPE and displays its bioactivity in an autocrine and paracrine fashion. The purpose of this study was to determine whether photoreceptors have the ability to synthesize NPD1, and whether or not this lipid mediator exerts bioactivity on these cells. For this purpose, 661W cells (mouse-derived photoreceptor cells) were used. First we asked whether these cells have the ability to form NPD1 by incubating cells with deuterium (d4)-labeled DHA exposed to dark and bright light treatments, followed by LC–MS/MS-based lipidomic analysis to identify and quantify d4-NPD1. The second question pertains to the potential bioactivity of these lipids. Therefore, cells were incubated with 9-cis-retinal in the presence of bright light that triggers cell damage and death. Following 9-cis-retinal loading, DHA, NPD1, or vehicle were added to the media and the 661W cells maintained either in darkness or under bright light. DHA and NPD1 were then quantified in cells and media. Regardless of lighting conditions, 661W cells acquired DHA from the media and synthesized 4–9 times as much d4-NPD1 under bright light treatment in the absence and presence of 9-cis-retinal compared to cells in darkness. Viability assays of 9-cis-retinal-treated cells demonstrated that 34 % of the cells survived without DHA or NPD1. However, after bright light exposure, DHA protected 23 % above control levels and NPD1 increased protection by 32 %. In conclusion, the photoreceptor cell line 661W has the capability to synthesize NPD1 from DHA when under stress, and, in turn, can be protected from stress-induced apoptosis by DHA or NPD1, indicating that photoreceptors effectively contribute to endogenous protective signaling mediated by NPD1 under stressful conditions.     

Mitochondrial Respiratory Dysfunction and Oxidative Stress after Chronic Malathion Exposure

Malathion is a pesticide used on a large scale and with high potential risk for human exposure. However, it is reasonable to hypothesize that while the malathion is metabolizing reactive oxygen species (ROS) can be generated and subsequently there is onset of an oxidative stress in central nervous system (CNS) structures: hippocampus, cortex, striatum and cerebellum of intoxicated rats due to mitochondrial respiratory chain disfunctions. The present study was therefore undertaken to evaluate malathion-induced lipid peroxidation (LPO), superoxide production from sub-mitochondrial particles and the activity of complexes II and IV of the mitochondrial respiratory chain. Malathion was administered in doses of 25, 50, 100 and 150 mg malathion/kg. After malathion administration LPO increased in hippocampus and striatum. This was accompanied by an increase in the formation of superoxide in submitochondrial particles in the hippocampus. Complex IV suffered significant inhibition of its activity. We could demonstrate in this study that malathion induces oxidative stress and it could be due to inactivation of mitochondrial respiratory complexes.     

Impaired Mitochondrial Dynamics and Nrf2 Signaling Contribute to Compromised Responses to Oxidative Stress in Striatal Cells Expressing Full-Length Mutant Huntingtin

Huntington disease (HD) is an inherited neurodegenerative disease resulting from an abnormal expansion of polyglutamine in huntingtin (Htt). Compromised oxidative stress defense systems have emerged as a contributing factor to the pathogenesis of HD. Indeed activation of the Nrf2 pathway, which plays a prominent role in mediating antioxidant responses, has been considered as a therapeutic strategy for the treatment of HD. Given the fact that there is an interrelationship between impairments in mitochondrial dynamics and increased oxidative stress, in this present study we examined the effect of mutant Htt (mHtt) on these two parameters. STHdh^Q111/Q111 cells, striatal cells expressing mHtt, display more fragmented mitochondria compared to STHdh^Q7/Q7 cells, striatal cells expressing wild type Htt, concurrent with alterations in the expression levels of Drp1 and Opa1, key regulators of mitochondrial fission and fusion, respectively. Studies of mitochondrial dynamics using cell fusion and mitochondrial targeted photo-switchable Dendra revealed that mitochondrial fusion is significantly decreased in STHdh^Q111/Q111 cells. Oxidative stress leads to dramatic increases in the number of STHdh^Q111/Q111 cells containing swollen mitochondria, while STHdh^Q7/Q7 cells just show increases in the number of fragmented mitochondria. mHtt expression results in reduced activity of Nrf2, and activation of the Nrf2 pathway by the oxidant tBHQ is significantly impaired in STHdh^Q111/Q111 cells. Nrf2 expression does not differ between the two cell types, but STHdh^Q111/Q111 cells show reduced expression of Keap1 and p62, key modulators of Nrf2 signaling. In addition, STHdh^Q111/Q111 cells exhibit increases in autophagy, whereas the basal level of autophagy activation is low in STHdh^Q7/Q7 cells. These results suggest that mHtt disrupts Nrf2 signaling which contributes to impaired mitochondrial dynamics and may enhance susceptibility to oxidative stress in STHdh^Q111/Q111 cells.     

Photo-Induced Oxidative Stress Impairs Mitochondrial Metabolism in Neurons and Astrocytes

Photodynamic therapy is selective destruction of cells stained with a photosensitizer upon irradiation with light at a specific wavelength in the presence of oxygen. Cell death upon photodynamic treatment is known to occur mainly due to free radical production and subsequent development of oxidative stress. During photodynamic therapy of brain tumors, healthy cells are also damaged; considering this, it is important to investigate the effect of the treatment on normal neurons and glia. We employed live-cell imaging technique to investigate the cellular mechanism of photodynamic action of radachlorin (200 nM) on neurons and astrocytes in primary rat cell culture. We found that the photodynamic effect of radachlorin increases production of reactive oxygen species measured by dihydroethidium and significantly decrease mitochondrial membrane potential. Mitochondrial depolarization was independent of opening of mitochondrial permeability transition pore and was insensitive to blocker of this pore cyclosporine A. However, irradiation of cells with radachlorin dramatically decreased NADH autofluorescence and also reduced mitochondrial NADH pool suggesting inhibition of mitochondrial respiration by limitation of substrate. This effect could be prevented by inhibition of poly (ADP-ribose) polymerase (PARP) with DPQ. Thus, irradiation of neurons and astrocytes in the presence of radachlorin leads to activation of PARP and decrease in NADH that leads to mitochondrial dysfunction.     

Arginase 2 Deficiency Prevents Oxidative Stress and Limits Hyperoxia-Induced Retinal Vascular Degeneration

Background

Hyperoxia exposure of premature infants causes obliteration of the immature retinal microvessels, leading to a condition of proliferative vitreoretinal neovascularization termed retinopathy of prematurity (ROP). Previous work has demonstrated that the hyperoxia-induced vascular injury is mediated by dysfunction of endothelial nitric oxide synthase resulting in peroxynitrite formation. This study was undertaken to determine the involvement of the ureahydrolase enzyme arginase in this pathology.

Methods and Findings

Studies were performed using hyperoxia-treated bovine retinal endothelial cells (BRE) and mice with oxygen-induced retinopathy (OIR) as experimental models of ROP. Treatment with the specific arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH) prevented hyperoxia-induced apoptosis of BRE cells and reduced vaso-obliteration in the OIR model. Furthermore, deletion of the arginase 2 gene protected against hyperoxia-induced vaso-obliteration, enhanced physiological vascular repair, and reduced retinal neovascularization in the OIR model. Additional deletion of one copy of arginase 1 did not improve the vascular pathology. Analyses of peroxynitrite by quantitation of its biomarker nitrotyrosine, superoxide by dihydroethidium imaging and NO formation by diaminofluoroscein imaging showed that the protective actions of arginase 2 deletion were associated with blockade of superoxide and peroxynitrite formation and normalization of NOS activity.

Conclusions

Our data demonstrate the involvement of arginase activity and arginase 2 expression in hyperoxia-induced vascular injury. Arginase 2 deletion prevents hyperoxia-induced retinal vascular injury by preventing NOS uncoupling resulting in decreased reactive oxygen species formation and increased nitric oxide bioavailability.     

氧化应激相关性疾病中线粒体机制的研究进展

氧化应激是由体内生成的活性氧（reactive oxygen species,ROS）／活性氮（reactivenitro．genspecies,RNS）与抗氧化防御机制之间的平衡被打破引起的,这与许多疾病的发病机理相关,包括神经退行性病变、肿瘤、炎症性疾病等。而线粒体作为细胞代谢的中枢,是其作用的主要靶细胞器,氧化应激引起线粒体内脂质、蛋白质与核酸的损伤,导致线粒体结构和功能的改变,该文就线粒体在上述与氧化应激相关的疾病中改变的研究进展作一综述。     

Protection Against Ischemic Brain Injury by Inhibition of Mitochondrial Oxidative Stress

Mitochondria are both targets and sources of oxidative stress. This dual relationship is particularly evident in experimental paradigms modeling ischemic brain injury. One mitochondrial metabolic enzyme that is particularly sensitive to oxidative inactivation is pyruvate dehydrogenase. This reaction is extremely important in the adult CNS that relies very heavily on carbohydrate metabolism, as it represents the sole bridge between anaerobic and aerobic metabolism. Oxidative injury to this enzyme and to other metabolic enzymes proximal to the electron transport chain may be responsible for the oxidized shift in cellular redox state that is observed during approximately the first hour of cerebral reperfusion. In addition to impairing cerebral energy metabolism, oxidative stress is a potent activator of apoptosis. The mechanisms responsible for this activation are poorly understood but likely involve the expression of p53 and possibly direct effects of reactive oxygen species on mitochondrial membrane proteins and lipids. Mitochondria also normally generate reactive oxygen species and contribute significantly to the elevated net production of these destructive agents during reperfusion. Approaches to inhibiting pathologic mitochondrial generation of reactive oxygen species include mild uncoupling, pharmacologic inhibition of the membrane permeability transition, and simply lowering the concentration of inspired oxygen. Antideath mitochondrial proteins of the Bcl-2 family also confer cellular resistance to oxidative stress, paradoxically through stimulation of mitochondrial free radical generation and secondary upregulation of antioxidant gene expression.     

Impaired Cellular Bioenergetics Causes Mitochondrial Calcium Handling Defects in MT-ND5 Mutant Cybrids

Mutations in mitochondrial DNA (mtDNA) can cause mitochondrial disease, a group of metabolic disorders that affect both children and adults. Interestingly, individual mtDNA mutations can cause very different clinical symptoms, however the factors that determine these phenotypes remain obscure. Defects in mitochondrial oxidative phosphorylation can disrupt cell signaling pathways, which may shape these disease phenotypes. In particular, mitochondria participate closely in cellular calcium signaling, with profound impact on cell function. Here, we examined the effects of a homoplasmic m.13565C>T mutation in MT-ND5 on cellular calcium handling using transmitochondrial cybrids (ND5 mutant cybrids). We found that the oxidation of NADH and mitochondrial membrane potential (Δψ_m) were significantly reduced in ND5 mutant cybrids. These metabolic defects were associated with a significant decrease in calcium uptake by ND5 mutant mitochondria in response to a calcium transient. Inhibition of glycolysis with 2-deoxy-D-glucose did not affect cytosolic calcium levels in control cybrids, but caused an increase in cytosolic calcium in ND5 mutant cybrids. This suggests that glycolytically-generated ATP is required not only to maintain Δψ_m in ND5 mutant mitochondria but is also critical for regulating cellular calcium homeostasis. We conclude that the m.13565C>T mutation in MT-ND5 causes defects in both mitochondrial oxidative metabolism and mitochondrial calcium sequestration. This disruption of mitochondrial calcium handling, which leads to defects in cellular calcium homeostasis, may be an important contributor to mitochondrial disease pathogenesis.     

Mitochondrial Alterations and Oxidative Stress in an Acute Transient Mouse Model of Muscle Degeneration: IMPLICATIONS FOR MUSCULAR DYSTROPHY AND RELATED MUSCLE PATHOLOGIES*

Muscular dystrophies (MDs) and inflammatory myopathies (IMs) are debilitating skeletal muscle disorders characterized by common pathological events including myodegeneration and inflammation. However, an experimental model representing both muscle pathologies and displaying most of the distinctive markers has not been characterized. We investigated the cardiotoxin (CTX)-mediated transient acute mouse model of muscle degeneration and compared the cardinal features with human MDs and IMs. The CTX model displayed degeneration, apoptosis, inflammation, loss of sarcolemmal complexes, sarcolemmal disruption, and ultrastructural changes characteristic of human MDs and IMs. Cell death caused by CTX involved calcium influx and mitochondrial damage both in murine C2C12 muscle cells and in mice. Mitochondrial proteomic analysis at the initial phase of degeneration in the model detected lowered expression of 80 mitochondrial proteins including subunits of respiratory complexes, ATP machinery, fatty acid metabolism, and Krebs cycle, which further decreased in expression during the peak degenerative phase. The mass spectrometry (MS) data were supported by enzyme assays, Western blot, and histochemistry. The CTX model also displayed markers of oxidative stress and a lowered glutathione reduced/oxidized ratio (GSH/GSSG) similar to MDs, human myopathies, and neurogenic atrophies. MS analysis identified 6 unique oxidized proteins from Duchenne muscular dystrophy samples (n = 6) (versus controls; n = 6), including two mitochondrial proteins. Interestingly, these mitochondrial proteins were down-regulated in the CTX model thereby linking oxidative stress and mitochondrial dysfunction. We conclude that mitochondrial alterations and oxidative damage significantly contribute to CTX-mediated muscle pathology with implications for human muscle diseases.     

Accumulation of Rhodopsin in Late Endosomes Triggers Photoreceptor Cell Degeneration

Progressive retinal degeneration is the underlying feature of many human retinal dystrophies. Previous work using Drosophila as a model system and analysis of specific mutations in human rhodopsin have uncovered a connection between rhodopsin endocytosis and retinal degeneration. In these mutants, rhodopsin and its regulatory protein arrestin form stable complexes, and endocytosis of these complexes causes photoreceptor cell death. In this study we show that the internalized rhodopsin is not degraded in the lysosome but instead accumulates in the late endosomes. Using mutants that are defective in late endosome to lysosome trafficking, we were able to show that rhodopsin accumulates in endosomal compartments in these mutants and leads to light-dependent retinal degeneration. Moreover, we also show that in dying photoreceptors the internalized rhodopsin is not degraded but instead shows characteristics of insoluble proteins. Together these data implicate buildup of rhodopsin in the late endosomal system as a novel trigger of death of photoreceptor neurons.     

Brucellosis Causes Alteration in Trace Elements and Oxidative Stress Factors

Brucellosis is regarded as one of the most common diseases among humans and livestock. In the present study, we aimed to assess the effect of this disease on the level of various cations including copper (Cu), manganese (Mn), zinc (Zn), and magnesium (Mg) as well as oxidative stress status in the serum of people suffering from brucellosis. The present case-study was carried out on 40 patients with brucellosis (case) and 20 healthy people (control). Blood specimens were taken from all the people and the level of essential trace elements and oxidative stress status were measured. The serum level of copper in the case group (165.39 ± 43.19 μg/dl) was significantly higher compared with that in the control group (122.12 ± 28.88 μg/dl). Whereas the serum level of zinc was significantly lower in the case group compared with that in the control group (76.47 ± 28.88 vs. 92.85 ± 23.16 μg/dl). The manganese and magnesium serum levels did not differ significantly between the two groups. Furthermore, total antioxidant capacity level was significantly lower in the case group (122.12 ± 28.22 μmol/ml) than that in the control group (3.08 ± 0.12 μmol/ml) and the level of serum malondialdehyde was significantly higher in the case group (7.20 ± 0.23 mmol/ml) than that in the control group (4.0 ± 0.19 mmol/ml). Brucellosis can cause alteration in the serum level of essential trace elements. Moreover, the present study indicated that brucellosis produces oxidative stress in patients.     

Altered Mitochondrial Function and Oxidative Stress in Leukocytes of Anorexia Nervosa Patients

Context

Anorexia nervosa is a common illness among adolescents and is characterised by oxidative stress.

Objective

The effects of anorexia on mitochondrial function and redox state in leukocytes from anorexic subjects were evaluated.

Design and setting

A multi-centre, cross-sectional case-control study was performed.

Patients

Our study population consisted of 20 anorexic patients and 20 age-matched controls, all of which were Caucasian women.

Main outcome measures

Anthropometric and metabolic parameters were evaluated in the study population. To assess whether anorexia nervosa affects mitochondrial function and redox state in leukocytes of anorexic patients, we measured mitochondrial oxygen consumption, membrane potential, reactive oxygen species production, glutathione levels, mitochondrial mass, and complex I and III activity in polymorphonuclear cells.

Results

Mitochondrial function was impaired in the leukocytes of the anorexic patients. This was evident in a decrease in mitochondrial O₂ consumption (P<0.05), mitochondrial membrane potential (P<0.01) and GSH levels (P<0.05), and an increase in ROS production (P<0.05) with respect to control subjects. Furthermore, a reduction of mitochondrial mass was detected in leukocytes of the anorexic patients (P<0.05), while the activity of mitochondrial complex I (P<0.001), but not that of complex III, was found to be inhibited in the same population.

Conclusions

Oxidative stress is produced in the leukocytes of anorexic patients and is closely related to mitochondrial dysfunction. Our results lead us to propose that the oxidative stress that occurs in anorexia takes place at mitochondrial complex I. Future research concerning mitochondrial dysfunction and oxidative stress should aim to determine the physiological mechanism involved in this effect and the physiological impact of anorexia.