Role of Mitochondrial Reactive Oxygen Species in the Activation of Cellular Signals, Molecules, and Function.

Mitochondria are a major source of intracellular energy and reactive oxygen species in cells, but are also increasingly being recognized as a controller of cell death. Here, we review evidence of signal transduction control by mitochondrial superoxide generation via the nuclear factor-κB (NF-κB) and GATA signaling pathways. We have also reviewed the effects of ROS on the activation of MMP and HIF. There is significant evidence to support the hypothesis that mitochondrial superoxide can initiate signaling pathways following transport into the cytosol. In this study, we provide evidence of TATA signal transductions by mitochondrial superoxide. Oxidative phosphorylation via the electron transfer chain, glycolysis, and generation of superoxide from mitochondria could be important factors in regulating signal transduction, cellular homeostasis, and cell death.

Chasing great paths of Helmut Sies "Oxidative Stress".

Prof. Dr. Helmut Sies is a pioneer of "Oxidative Stress", and has published over 18 papers with the name of "Oxidative Stress" in the title. He has been Editor-in-Chief of the journal "Archives of Biochemistry and Biophysics" for many years, and is a former Editor-in-Chief of the journal "Free Radical Research". He has clarified our understanding of the causes of chronic developing diseases, and has studied antioxidant factors. In this article, importance of "Oxidative Stress" and our mitochondrial oxidative stress studies; roles of mitochondrial ROS, effects of vitamin E and its homologues in oxidative stress-related diseases, effects of antioxidants in vivo and in vitro, and a mitochondrial superoxide theory for oxidative stress diseases and aging are introduced, and some of our interactions with Helmut are described, congratulating and appreciating his great path.

MnSOD downregulation induced by extremely low 0.1 mGy single and fractionated X-rays and microgravity treatment in human neuroblastoma cell line, NB-1.

A human neuroblastoma cell line, NB-1, was treated with 24 h of microgravity simulation by clinostat, or irradiated with extremely small X-ray doses of 0.1 or 1.0 mGy using single and 10 times fractionation regimes with 1 and 2 h time-intervals. A quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) examination was performed for apoptosis related factors (BAX, CYTC, APAF1, VDAC1-3, CASP3, CASP8, CASP9 P53, AIF, ANT1 and 2, BCL2, MnSOD, autophagy related BECN and necrosis related CYP-40. The qRT-PCR results revealed that microgravity did not result in significant changes except for a upregulation of proapoptotic VDAC2, and downregulations of proapoptotic CASP9 and antiapoptotic MnSOD. After 0.1 mGy fractionation irradiation, there was increased expression of proapoptotic APAF1 and downregulation of proapoptotic CYTC, VDAC2, VDAC3, CASP8, AIF, ANT1, and ANT2, as well as an increase in expression of antiapoptotic BCL2. There was also a decrease in MnSOD expression with 0.1 mGy fractionation irradiation. These results suggest that microgravity and low-dose radiation may decrease apoptosis but may potentially increase oxidative stress.

A mitochondrial superoxide theory for oxidative stress diseases and aging.

Fridovich identified CuZnSOD in 1969 and manganese superoxide dismutase (MnSOD) in 1973, and proposed "the Superoxide Theory," which postulates that superoxide (O2 (•-)) is the origin of most reactive oxygen species (ROS) and that it undergoes a chain reaction in a cell, playing a central role in the ROS producing system. Increased oxidative stress on an organism causes damage to cells, the smallest constituent unit of an organism, which can lead to the onset of a variety of chronic diseases, such as Alzheimer's, Parkinson's, amyotrophic lateral sclerosis and other neurological diseases caused by abnormalities in biological defenses or increased intracellular reactive oxygen levels. Oxidative stress also plays a role in aging. Antioxidant systems, including non-enzyme low-molecular-weight antioxidants (such as, vitamins A, C and E, polyphenols, glutathione, and coenzyme Q10) and antioxidant enzymes, fight against oxidants in cells. Superoxide is considered to be a major factor in oxidant toxicity, and mitochondrial MnSOD enzymes constitute an essential defense against superoxide. Mitochondria are the major source of superoxide. The reaction of superoxide generated from mitochondria with nitric oxide is faster than SOD catalyzed reaction, and produces peroxynitrite. Thus, based on research conducted after Fridovich's seminal studies, we now propose a modified superoxide theory; i.e., superoxide is the origin of reactive oxygen and nitrogen species (RONS) and, as such, causes various redox related diseases and aging.

Condylar shape in relation to anterior disk displacement in juvenile females.

The purpose of the study was to test the hypothesis that condylar shape varies based upon the condition of anterior disk displacement in young adolescent patients with temporomandibular disorder (TMD). The study design consisted of 96 juvenile female patients (aged 9 to 20; 15.1 +/- 2.3 yrs.) with clinical signs and/or symptoms of TMD. Bilateral high-resolution magnetic resonance imaging scans were performed in frontal and horizontal views with the mandible in the closed position. Disk positions were evaluated to classify the patients into three diagnostic groups. The results of the study, using ANOVA and Bonferroni tests, demonstrated significant differences among the groups. The conclusion drawn from the study was that condylar shape and size vary based on anterior disk position in juvenile females with TMD. The study's results suggest that disk displacement results in a smaller condyle.

Clinical significance of magnetization transfer contrast imaging for edematous changes in masticatory muscle.

OBJECTIVE: To evaluate inflammatory changes in masticatory muscles by magnetization transfer contrast (MTC) imaging. METHODS: Gradient-recalled echo (GRE) and MTC-GRE images of relaxed masticatory muscles in 28 healthy volunteers were obtained before and after exercise. At the same time, muscle stiffness and pain in the masseter muscles were also measured. Magnetization transfer ratios (MTRs) of the muscles were calculated from the GRE and MTC-GRE images. The MTRs of the masticatory muscles in 50 patients with temporomandibular disorder were compared with those in the volunteers. RESULTS: Immediately after the exercise, the MTRs of the masseter muscles significantly decreased (P < 0.05), whereas muscle stiffness and pain increased in the healthy volunteers. In patients with masseter muscle pain, the MTRs of the masseter muscles were significantly lower than in the healthy volunteers (P < 0.05). CONCLUSIONS: Magnetization transfer contrast imaging strongly reflects the masticatory muscle edematous changes, possibly leading to masseter muscle pain.

Author Correction: Changes in mitochondrial homeostasis and redox status in astronauts following long stays in space.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Clinical guidelines for dental cone-beam computed tomography.

Dental cone-beam computed tomography (CBCT) received regulatory approval in Japan in 2000 and has been widely used since being approved for coverage by the National Health Insurance system in 2012. This imaging technique allows dental practitioners to observe and diagnose lesions in the dental hard tissue in three dimensions (3D). When performing routine radiography, the examination must be justified, and optimal protection should be provided according to the ALARA (as low as reasonably achievable) principles laid down by the International Commission on Radiological Protection. Dental CBCT should be performed in such a way that the radiation exposure is minimized and the benefits to the patient are maximized. There is a growing demand for widespread access to cutting-edge health care through Japan's universal health insurance system. However, at the same time, people want our limited human, material, and financial resources to be used efficiently while providing safe health care at the least possible cost to society. Japan's aging population is expected to reach a peak in 2025, when most of the baby boomer generation will be aged 75 years or older. Comprehensive health care networks are needed to overcome these challenges. Against this background, we hope that this text will contribute to the nation's oral health by encouraging efficient use of dental CBCT.

Evidence of ROS generation by mitochondria in cells with impaired electron transport chain and mitochondrial DNA damage.

Mitochondrial damage is a well known cause of mitochondria-related diseases. A major mechanism underlying the development of mitochondria-related diseases is thought to be an increase in intracellular oxidative stress produced by impairment of the mitochondrial electron transport chain (ETC). However, clear evidence of intracellular free radical generation has not been clearly provided for mitochondrial DNA (mtDNA)-damaged cells. In this study, using the novel fluorescence dye, 2-[6-(4'-hydroxy)phenoxy-3H-xanthen-3-on-9-yl]benzoic acid (HPF), which was designed to detect hydroxyl radicals (*OH), intracellular free radical formation was examined in 143B cells (parental cells), 143B-rho(0) cells (mtDNA-lacking cells), 87 wt (cybrid), and cybrids of 4977-bp mtDNA deletion (common deletion) cells containing the deletion with 0%, 5%, 50% and >99% frequency (HeLacot, BH5, BH50 and BH3.12, respectively), using a laser confocal microscope detection method. ETC inhibitors (rotenone, 3-nitropropionic acid, thenoyltrifluoroacetone, antimycin A and sodium cyanide) were also tested to determine whether inhibitor treatment increased intracellular reactive oxygen species (ROS) generation. A significant increase in ROS for 143B-rho(0) cells was observed compared with 143B cells. However, for the 87 wt cybrid, no increase was observed. An increase was also observed in the mtDNA-deleted cells BH50 and BH3.12. The ETC inhibitors increased intracellular ROS in both 143B and 143B-rho(0) cells. Furthermore, in every fluorescence image, the fluorescence dye appeared localized around the nuclei. To clarify the localization, we double-stained cells with the dye and MitoTracker Red. The resulting fluorescence was consistently located in mitochondria. Furthermore, manganese superoxide dismutase (MnSOD) cDNA-transfected cells had decreased ROS. These results suggest that more ROS are generated from mitochondria in ETC-inhibited and mtDNA-damaged cells, which have impaired ETC.

The usefulness of diagnostic imaging for the assessment of pain symptoms in temporomandibular disorders.

The causes of pain symptoms in the temporomandibular joint (TMJ) and masticatory muscle (MM) regions may not be determined by clinical examination alone. In this review, we document that pain symptoms of the TMJ and MM regions in patients with temporomandibular disorders (TMDs) are associated with computed tomography and magnetic resonance (MR) findings of internal derangement, joint effusion, osteoarthritis, and bone marrow edema. However, it is emphasized that these imaging findings must not be regarded as the unique and dominant factors in defining TMJ pain. High signal intensity and prominent enhancement of the posterior disk attachment on fat saturation T2-weighted imaging and dynamic MR imaging with contrast material are closely correlated with the severity of TMJ pain. Magnetic transfer contrast, MR spectroscopy, diffusion tensor imaging, and ultrasonography findings have helped identify intramuscular edema and contracture as one of the causes of MM pain and fatigue. Recently, changes in brain as detected by functional MR neuroimaging have been associated with changes in the TMJ and MM regions. The thalamus, the primary somatosensory cortex, the insula, and the anterior and mid-cinglate cortices are most frequently associated with TMD pain.

Changes in mitochondrial homeostasis and redox status in astronauts following long stays in space.

The effects of long-term exposure to extreme space conditions on astronauts were investigated by analyzing hair samples from ten astronauts who had spent six months on the International Space Station (ISS). Two samples were collected before, during and after their stays in the ISS; hereafter, referred to as Preflight, Inflight and Postflight, respectively. The ratios of mitochondrial (mt) to nuclear (n) DNA and mtRNA to nRNA were analyzed via quantitative PCR. The combined data of Preflight, Inflight and Postflight show a significant reduction in the mtDNA/nDNA in Inflight, and significant reductions in the mtRNA/nRNA ratios in both the Inflight and Postflight samples. The mtRNA/mtDNA ratios were relatively constant, except in the Postflight samples. Using the same samples, the expression of redox and signal transduction related genes, MnSOD, CuZnSOD, Nrf2, Keap1, GPx4 and Catalase was also examined. The results of the combined data from Preflight, Inflight and Postflight show a significant decrease in the expression of all of the redox-related genes in the samples collected Postflight, with the exception of Catalase, which show no change. This decreased expression may contribute to increased oxidative stress Inflight resulting in the mitochondrial damage that is apparent Postflight.

Roles of mitochondria-generated reactive oxygen species on X-ray-induced apoptosis in a human hepatocellular carcinoma cell line, HLE.

HLE, a human hepatocellular carcinoma cell line was transiently transfected with normal human MnSOD and MnSOD without a mitochondrial targeting signal (MTS). Mitochondrial reactive oxygen species (ROS), lipid peroxidation and apoptosis were examined as a function of time following 18.8 Gy X-ray irradiation. Our results showed that the level of mitochondrial ROS increased and reached a maximum level 2 hours after X-ray irradiation. Authentic MnSOD, but not MnSOD lacking MTS, protected against mitochondrial ROS, lipid peroxidation and apoptosis. In addition, the levels of mitochondrial ROS were consistently found to always correlate with the levels of authentic MnSOD in mitochondria. These results suggest that only when MnSOD is located in mitochondria is it efficient in protecting against cellular injuries by X-ray irradiation and that mitochondria are the critical sites of X-ray-induced cellular oxidative injuries.

Mitochondria as possible pharmaceutical targets for the effects of vitamin E and its homologues in oxidative stress-related diseases.

It is well known that vitamin E functions as an antioxidant, and it is expected to exert an antioxidant effect when taken as a supplement. However, a number of cohort studies have shown that vitamin E does not alleviate oxidative stress and could even worsen it. Recently, Wang et al. investigated whether vitamin E intake was associated with amyotrophic lateral sclerosis (ALS) based on data from 5 cohort studies with 1,055,546 participants, of which 805 of them had developed ALS. They concluded in this large pooled prospective study, in which long-term vitamin E supplementation was associated with lower ALS rates, and therefore, a possible protective effect of vitamin E deserves further consideration. Performing further large cohort studies may reveal similar findings for other oxidative stress-related diseases. It is still controversial if antioxidants such as vitamin E provide a clinical therapeutic effect against oxidative stress-related diseases. If effective, the dose at which they should be administered and the duration of supplement exposure should be of interest. Vitamin E reduces production of reactive oxygen species by mitochondria and elicits further reactions in cells. It should be noted that mitochondria are important targets for vitamin E and its homologues. Therefore, a proper usage of vitamin E in subjects under high oxidative stress, due to its individually targeting property, will arise its importance in healthy life.