Mitochondrial Cell Research Articles

PO2-based biodosimetry evaluation using an EPR technique acts as a sensitive index for chemotherapy.

The partial pressure of oxygen (PO2) in the tumor microenvironment directly affects tumor sensitivity to chemotherapy. In the present study, a lithium phthalocyanine probe was implanted into MCF-7 human breast cancer cells, followed by transplant of the cells into nude mice. The present study used an electron paramagnetic resonance (EPR) oximetry measuring technique to dynamically monitor PO2 in the tumor microenvironment prior to and following chemotherapy, and aimed to determine the precise time window in which the microenvironmental PO2 peaked following chemotherapy. The results indicated that PO2 was significantly higher in breast cancer compared with control (P<0.05). Following four cycles of chemotherapy, the activity of NADH dehydrogenase, succinate-cytochrome c reductase and cytochrome c oxidase in the mitochondria of cells was significantly reduced when compared with their activity prior to chemotherapy (P<0.05). Regional blood flow in tumor tissues undergoing chemotherapy was significantly lower than that prior to chemotherapy (P<0.05). The rate of cellular apoptosis in the PO2 peak-based chemotherapy group was significantly greater than that in the conventional chemotherapy group after two and four cycles of chemotherapy (P<0.05). Tumor volume in the PO2 peak-based chemotherapy group was significantly reduced compared with that in the 0.9% NaCl solution control and the conventional chemotherapy groups after four cycles of chemotherapy (P<0.05). The tumor inhibitory rate of the experimental group was significantly higher than that of the conventional chemotherapy group (P<0.01). In conclusion, the present study may provide guidance for the development of effective strategies depending on tumor-maximal response to chemotherapy in an oxygen-rich environment. Additionally, the present study aimed to establish a foundation for a clinical noninvasive assessment intended to guide treatment and formulate individual regimens, in order to improve cancer therapeutics, sensitivity monitoring and curative effect estimation.

Mitochondrial HSP60 Protein in Rat Medulla Oblongata Cells

HSP60 is a member of the heat shock protein (HSP) group and is located predominantly in cell mitochondria. Dysfunction and insufficient synthesis of HSP60 in the nervous system leads to neurodegeneration. The aim of the present work was to conduct immunohistochemical determination of the location of HSP60 protein in neurons and gliocytes in the medulla oblongata in rats. Studies were performed on mature male Wistar rats weighing 200–250 g (n = 7). The results provided evidence that HSP60 has a discrete localization in cytoplasmic structures in neurons and gliocytes. In large neurons, local accumulation of HSP60 allows fine structures to be detected, probably mitochondria. An important observation is that of the absence of any cytosolic reaction in medulla oblongata neurons and gliocytes. This provides grounds for using the reaction for HSP60 in complex studies of apoptosis in nerve cells, as this is associated with an accumulation of HSP60 and displacement of immunopositive product to the plasmalemma.

Host–Guest Complexation of a Lipoic Acid Conjugate of Calix[4]arene with Pyridinium Moiety on Gold Nanorods for Mitochondrial Tracking Followed by Cytotoxicity in HeLa Cells under 633 nm Laser Light

A new 1,3-lower rim-lipoic acid appended calix[4]arene (L) has been synthesized and characterized by different spectral techniques. Lipoic acid functionality helps L to anchor onto a noble metal surface, and in the present case these are the gold nanorods (GNRs). As the conjugate of calix[4]arene acts as a host due to the presence of its arene cavity, pyridinium containing guests have been explored to study their complexation, since the pyridinium containing molecules will accumulate in mitochondria in cells. The host–guest complexation has been explored in solution by 1H NMR, ESI-MS, UV–vis, and fluorescence spectral titration studies. The loading of host and the guest onto the GNR surface has been analyzed by spectroscopy and microscopy and found that their combination results in nanocomplexation. The presence of pyridinium functionality on the guest leads to targeted delivery to mitochondria of GNR nanocomposites as shown by confocal laser scanning microscopy imaging. The nanocomplex has been studied f...

Carboxylate Analogues of Aryl-Urea-Substituted Fatty Acids That Target the Mitochondria in MDA-MB-231 Breast Cancer Cells to Promote Cell Death.

Selective targeting of the tumor cell mitochondrion is a viable approach for the development of anticancer agents because the organelle is functionally different from the mitochondria of normal cells. We recently developed a novel aryl-urea fatty acid, 16({[4-chloro-3-(trifluoromethyl)phenyl]carbamoyl}amino)hexadecanoic acid (1) that was found to disrupt mitochondria and to activate apoptosis in MDA-MB-231 breast cancer cells. However, there is currently little information on the structural requirements for the activity of compound 1 analogues. The present study evaluated the role of the carboxylic acid group on the anticancer activity of 1. Bioisosteric replacement of the carboxylate in 1 maintained activity. Thus, like 1, the sulfonic acid analogue 1-SA and the oxo-thiadiazole analogue 1-OT were also found to target the mitochondrion and to activate cell killing capacity. The hydroxamic acid analogue 1-HA also killed MDA-MB-231 cells, but its onset of action was slower than that of 1-SA and 1-OT. In contrast, replacement of the carboxylate with non-bioisosteric amido and methylamido groups produced analogues that minimally altered mitochondrial function and showed little capacity to decrease tumor cell viability. These findings suggest that the carboxylate moiety in the novel mitochondrially targeted agent 1 is an important determinant of the kinetics and efficacy of anticancer cell activities of compound 1 analogues. Further development of carboxylate-modified analogues of aryl-urea fatty acids as potential anticancer agents could now be warranted.

18.1 MITOCHONDRIAL ALTERATIONS WITHIN THE PYRAMIDAL-PARVALBUMIN CELL MICROCIRCUIT IN THE PREFRONTAL CORTEX IN SCHIZOPHRENIA

BackgroundWorking memory, a core cognitive function impaired in schizophrenia, depends upon gamma oscillatory neuronal activity in the prefrontal cortex (PFC). Accordingly, individuals with schizophrenia show lower power of gamma oscillations in the PFC during tasks that involve working memory. Gamma oscillations emerge from the fast and coordinated activity of layer 3 excitatory pyramidal cells and inhibitory parvalbumin (PV) cells. As such, gamma oscillations have a particularly high energetic demand that is met by ATP production via oxidative phosphorylation (OXPHOS) within pyramidal and PV cell mitochondria. PFC layer 3 pyramidal cells have prominent reductions in OXPHOS-related gene pathways in schizophrenia. Importantly, OXPHOS can be regulated by two distinct processes: ATP demand to support neuronal firing, or upstream deficits in OXPHOS enzyme expression. Layer- and cell type-specific transcriptomic analyses of OXPHOS enzymes and ultrastructural analyses of mitochondrial morphology can help to distinguish between these two possibilities. Reduced ATP demand due to reduced neuronal firing is associated with 1) correlated expression levels of OXPHOS enzyme complexes, 2) lower expression of all subunits comprising Complex IV, the terminal and rate-limiting OXPHOS enzyme complex, and 3) normal mitochondrial morphology. Defective OXPHOS results in 1) elimination of correlated expression of OXPHOS enzyme complexes, 2) variable and inconsistent effects on Complex IV subunit expression, and 3) abnormal mitochondrial morphology.To determine which upstream factor is likely operative in the illness, we quantified OXPHOS enzyme complex transcripts in layer 3 pyramidal and PV cells, and performed ultrastructural analyses of mitochondrial morphology within pyramidal and PV axon boutons in layer 3 of the PFC in schizophrenia and unaffected comparison subjects.MethodsFor mRNA analyses, frozen tissue sections of area 9 from 36 pairs of schizophrenia and comparison subjects were stained for Nissl substance to identify pyramidal cells, or labeled using immunoperoxidase for aggrecan to identify PV cells. Layer 3 pyramidal and PV somata were dissected using laser capture microdissection. Transcriptome profiling was performed by microarray using Affymetrix GeneChips. Analysis of Complexes I, IV, and V expression in each neuronal population was assessed at q<0.05 in covariate- and multiple comparisons-corrected analyses.Electron microscopic analyses were performed in area 46 of 2 matched pairs of schizophrenia and comparison subjects. Mitochondria in pyramidal and PV cell boutons were classified as normal or abnormal using established criteria. Chi-square analysis was used to examine whether the percentages of each type differed between groups.ResultsThe expression of subunits comprising Complexes I, IV, and V in layer 3 pyramidal and PV cells was significantly lower in subjects with schizophrenia relative to unaffected comparison subjects. In both cell populations, expression of Complexes I, IV, and V were correlated (r=0.8-0.9) in unaffected comparison and schizophrenia subjects. Complex IV subunits showed 11–26% reductions in pyramidal cells, and 7-31% reductions in PV cells in schizophrenia. In both unaffected comparison and schizophrenia subjects, ≥99% of mitochondria in pyramidal cell boutons and ≥97% of mitochondria in PV cell boutons exhibited normal morphology.DiscussionThe current findings are most consistent with the interpretation that lower measures of OXPHOS in schizophrenia reflect lower demand for ATP production due to less neuronal firing.

Intraflagellar transport is deeply integrated in hedgehog signaling.

The vertebrate hedgehog pathway is organized in primary cilia, and hedgehog components relocate into or out of cilia during signaling. Defects in intraflagellar transport (IFT) typically disrupt ciliary assembly and attenuate hedgehog signaling. Determining whether IFT drives the movement of hedgehog components is difficult due to the requirement of IFT for building cilia. Unlike most IFT proteins, IFT27 is dispensable for cilia formation but affects hedgehog signaling similarly to other IFTs, allowing us to examine its role in the dynamics of signaling. Activating signaling at points along the pathway in Ift27 mutant cells showed that IFT is extensively involved in the pathway. Similar analysis of Bbs mutant cells showed that BBS proteins participate at many levels of signaling but are not needed to concentrate Gli transcription factors at the ciliary tip. Our analysis showed that smoothened delivery to cilia does not require IFT27, but the role of other IFTs is not known. Using a rapamycin-induced dimerization system to sequester IFT-B proteins at the mitochondria in cells with fully formed cilia did not affect the delivery of Smo to cilia, suggesting that this membrane protein may not require IFT-B for delivery.

Pro-fluorescent mitochondria-targeted real-time responsive redox probes synthesised from carboxy isoindoline nitroxides: Sensitive probes of mitochondrial redox status in cells

Here we describe new fluorescent probes based on fluorescein and rhodamine that provide reversible, real-time insight into cellular redox status. The new probes incorporate bio-imaging relevant fluorophores derived from fluorescein and rhodamine linked with stable nitroxide radicals such that they cannot be cleaved, either spontaneously or enzymatically by cellular processes. Overall fluorescence emission is determined by reversible reduction and oxidation, hence the steady state emission intensity reflects the balance between redox potentials of critical redox couples within the cell. The permanent positive charge on the rhodamine-based probes leads to their rapid localisation within mitochondria in cells. Reduction and oxidation also leads to marked changes in the fluorophore lifetime, enabling monitoring by fluorescence lifetime imaging microscopy. Finally, we demonstrate that administration of a methyl ester version of the rhodamine-based probe can be used at concentrations as low as 5 nM to generate a readily detected response to redox stress within cells as analysed by flow cytometry.

Mitochondria in smooth muscle cells of viscera.

The spatial density of mitochondria was studied by thin-section electron microscopy in smooth muscles of bladder, iris and gut in mice, rats, guinea-pigs and sheep. Morphometric data included areas of muscle cell profiles (~6,000 muscle cells were measured) and areas of their mitochondria (more than three times as many). The visual method delivers accurate estimates of the extent of the chondrioma (the ensemble of mitochondria in a cell), measuring all and only the mitochondria in each muscle cell and no other cells. The digital records obtained can be used again for checks and new searches. Spatial density of mitochondria varies between about 2 and 10% in different muscles in different species. In contrast, there is consistency of mitochondrial density within a given muscle in a given species. For each muscle in each species there is a characteristic mitochondrial density with modest variation between experiments. On the basis of data from serial sections in the rat detrusor muscle, mitochondrial density varies very little between the muscle cells, each cell having a value close to that for the whole muscle. Mitochondrial density is different in a given muscle, e.g., ileal circular muscle, from the four mammalian species, with highest values in mouse and lowest in sheep; in mice the mitochondrial density is nearly three time higher that in sheep. In a given species there are characteristic variations between different muscles. For example, the bladder detrusor muscle has markedly fewer mitochondria than the ileum, and the iris has markedly more.

Development of surface-engineered PLGA nanoparticulate-delivery system of Tet1-conjugated nattokinase enzyme for inhibition of Aβ40 plaques in Alzheimer's disease.

According to the World Health Organization, globally there are around 18 million patients suffering from Alzheimer’s disease (AD), and this number is expected to double by 2025. The pathophysiology of AD includes selective deposition of Aβ peptide in the mitochondria of cells, which inhibits uptake of glucose by neurons and key enzyme functions. Current drug treatments for AD are unable to rectify the underlying pathology of the disease; they only provide short-term symptomatic relief, so there is a need for the development of newer treatment regimes. The antiamyloid activity, antifibrinolytic activity, and antithrombotic activity of nattokinase holds potential for the treatment of AD. As nattokinase is a protein, its stability restricts its usage to a greater extent, but this limitation can be overcome by nanoencapsulation. In this work, we successfully synthesized polymeric nanoparticles of nattokinase and characterized its use by different techniques: transmission electron microscopy, scanning electron microscopy, DTS Nano, differential scanning calorimetry, Fourier-transform infrared spectroscopy, thioflavin T-binding assay, in vitro drug release, antifibrinolytic activity, and in vivo antiamyloid activity. As brain targeting of hydrophilic drugs is complicated due to the stringent nature of blood–brain barrier, in the current experimental study, we conjugated poly(lactic-co-glycolic acid) (PLGA)-encapsulated nattokinase with Tet1 peptide, which exhibits retrograde transportation properties because of its affinity to neurons. Our study suggests that PLGA-encapsulated nattokinase polymeric nanoparticles are able to downregulate amyloid aggregation and exhibit antifibrinolytic activity. The encapsulation of nattokinase in PLGA did not affect its enzyme activity, so the prepared nanoformulation containing nattokinase can be used as an effective drug treatment against AD.

Aggregation-Induced Fluorescence Probe for Monitoring Membrane Potential Changes in Mitochondria.

Fluorescent probe 2 was designed for selectively determining mitochondria membrane potential changes. The probe selectively detects changes in the mitochondria membrane potential in a manner that is more sensitive than that of the commercially available indicator, Rodamine 123. As a result, the probe 2 is ideal for future studies designed to assess the functions of mitochondria in cells.

Visualization of Multicolored in vivo Organelle Markers for Co-Localization Studies in Oryza sativa.

Eukaryotic cells consist of a complex network of thousands of proteins present in different organelles where organelle-specific cellular processes occur. Identification of the subcellular localization of a protein is important for understanding its potential biochemical functions. In the post-genomic era, localization of unknown proteins is achieved using multiple tools including a fluorescent-tagged protein approach. Several fluorescent-tagged protein organelle markers have been introduced into dicot plants, but its use is still limited in monocot plants. Here, we generated a set of multicolored organelle markers (fluorescent-tagged proteins) based on well-established targeting sequences. We used a series of pGWBs binary vectors to ameliorate localization and co-localization experiments using monocot plants. We constructed different fluorescent-tagged markers to visualize rice cell organelles, i.e., nucleus, plastids, mitochondria, peroxisomes, golgi body, endoplasmic reticulum, plasma membrane, and tonoplast, with four different fluorescent proteins (FPs) (G3GFP, mRFP, YFP, and CFP). Visualization of FP-tagged markers in their respective compartments has been reported for dicot and monocot plants. The comparative localization of the nucleus marker with a nucleus localizing sequence, and the similar, characteristic morphology of mCherry-tagged Arabidopsis organelle markers and our generated organelle markers in onion cells, provide further evidence for the correct subcellular localization of the Oryza sativa (rice) organelle marker. The set of eight different rice organelle markers with four different FPs provides a valuable resource for determining the sub-cellular localization of newly identified proteins, conducting co-localization assays, and generating stable transgenic localization in monocot plants.

Functional Mitochondria in Health and Disease.

The ability to rapidly adapt cellular bioenergetic capabilities to meet rapidly changing environmental conditions is mandatory for normal cellular function and for cancer progression. Any loss of this adaptive response has the potential to compromise cellular function and render the cell more susceptible to external stressors such as oxidative stress, radiation, chemotherapeutic drugs, and hypoxia. Mitochondria play a vital role in bioenergetic and biosynthetic pathways and can rapidly adjust to meet the metabolic needs of the cell. Increased demand is met by mitochondrial biogenesis and fusion of individual mitochondria into dynamic networks, whereas a decrease in demand results in the removal of superfluous mitochondria through fission and mitophagy. Effective communication between nucleus and mitochondria (mito-nuclear cross talk), involving the generation of different mitochondrial stress signals as well as the nuclear stress response pathways to deal with these stressors, maintains bioenergetic homeostasis under most conditions. However, when mitochondrial DNA (mtDNA) mutations accumulate and mito-nuclear cross talk falters, mitochondria fail to deliver critical functional outputs. Mutations in mtDNA have been implicated in neuromuscular and neurodegenerative mitochondriopathies and complex diseases such as diabetes, cardiovascular diseases, gastrointestinal disorders, skin disorders, aging, and cancer. In some cases, drastic measures such as acquisition of new mitochondria from donor cells occurs to ensure cell survival. This review starts with a brief discussion of the evolutionary origin of mitochondria and summarizes how mutations in mtDNA lead to mitochondriopathies and other degenerative diseases. Mito-nuclear cross talk, including various stress signals generated by mitochondria and corresponding stress response pathways activated by the nucleus are summarized. We also introduce and discuss a small family of recently discovered hormone-like mitopeptides that modulate body metabolism. Under conditions of severe mitochondrial stress, mitochondria have been shown to traffic between cells, replacing mitochondria in cells with damaged and malfunctional mtDNA. Understanding the processes involved in cellular bioenergetics and metabolic adaptation has the potential to generate new knowledge that will lead to improved treatment of many of the metabolic, degenerative, and age-related inflammatory diseases that characterize modern societies.

Membrane-bound ATP-dependent Energy Systems, as Extra- and Intracellular Key Signals for Gastrointestinal Functions and their Regulations in the Gastrointestinal Mucosa.

Our research group has carried out various biochemical examinations in rat gastric ulcer models and in human gastrointestinal resecates obtained from patient who underwent gastric intervention due to peptic ulcer disease. Biochemical methods gave excellent possibility to approach the biochemical events taking place in tissues, cellular and subcellular regulatory levels during of ulcer development and of its prevetions. This paper gives a brief summary of these biochemical examinations conducted during this study period started from the 1960's up till now. 1. The decreased action of gastric acid secretory responses is not needed for duodenal and gastric ulcer healing in patients with peptic ulcer; 2. The surgical and chemical vagotomy resulted in various biochemical changes in the rat stomach; 3. The presence of Na+-K+-dependent ATPase and adenylate cyclase can be demostrated both in the rat and human gastric fundic mucosa; 4. The mitocondrial ATP is a common substrate for these membrane-bound ATP-dependent enzymes, and a multiple feedback mechanism existing between these two membrane-bound enzymes altered by mediators, hormones and drugs; 5. This feedback mechanism exists in the GI mucosa under different pathological conditions and during certain drug actions; 6. The development of mucosal damage and prevention depends on the actual regulatory state of above mentioned feedback mechanism between the membrane-bound ATP-dependent energy systems; 7.The drug actions depend on the actual functional state of target organ; 8. Biochemical gradients exist between the biochemical structure of the fundic, antral, duodenal and jejunal mucosa in patients with gastric hyperacidity, which is gradually downregulated by the decrease of gastric acid secretory responses, and totally disappears in patients with hypacidity; 9. No biochemically proven tissue hypoxia - around the chronic ulcer, duodenal and jejunal ulcers - exists in patients with chronic peptic ulcer; 10. The cellular and tissue protection differ from each other in the gastrointestinal tract; 11. Helicobacter pylori does not produce damage at the level of cell membrane, mitochondrion and DNA - given alone or in combination with indomethacin - on freshly isolated rat gastric mucosal cells. 12. A biochemical explanation is given to ulcer development in humans and in different animal models.

Rifampicin inhibits rotenone-induced microglial inflammation via enhancement of autophagy

Mitochondrial and autophagic dysfunction, as well as neuroinflammation, are associated with the pathophysiology of Parkinson's disease (PD). Rotenone, an inhibitor of mitochondrial complex I, has been associated as an environmental neurotoxin related to PD. Our previous studies reported that rifampicin inhibited microglia activation and production of proinflammatory mediators induced by rotenone, but the precise mechanism has not been completely elucidated. BV2 cells were pretreated for 2h with rifampicin followed by 0.1μM rotenone, alone or in combination with chloroquine. Here, we demonstrate that rifampicin pretreatment alleviated rotenone induced release of IL-1β and IL-6, and its effects were suppressed when autophagy was inhibited by chloroquine. Moreover, preconditioning with 50μM rifampicin significantly increased viability of SH-SY5Y cells cocultured with rotenone-treated BV2 cells in the transwell coculture system. Chloroquine partially abolished the neuroprotective effects of rifampicin pretreatment. Rifampicin pretreatment significantly reversed rotenone-induced mitochondrial membrane potential reduction and reactive oxygen species accumulation. We suggest that the mechanism for rifampicin-mediated anti-inflammatory and antioxidant effects is the enhancement of autophagy. Indeed, the ratio of LC3-II/LC3-I in rifampicin-pretreated BV2 cells was significantly higher than that in cells without pretreatment. Fluorescence and electron microscopy analyses indicate an increase of lysosomes colocalized with mitochondria in cells pretreated with rifampicin, which confirms that the damaged mitochondria were cleared through autophagy (mitophagy). Taken together, the data provide further evidence that rifampicin exerts neuroprotection against rotenone-induced microglia inflammation, partially through the autophagy pathway. Modulation of autophagy by rifampicin is a novel therapeutic strategy for PD.

To mdivi-1 or not to mdivi-1: Is that the question?

The fission/division and fusion of mitochondria are fundamental aspects of mitochondrial biology. The balance of fission and fusion sets the length of mitochondria in cells to serve their physiological requirements. The fission of mitochondria is markedly induced in many disease states and in response to cellular injury, resulting in the fragmentation of mitochondria into dysfunctional units. The mechanism that drives fission is dependent on the dynamin related protein 1 (Drp1) GTPase. mdivi-1 is a quinazolinone originally described as a selective inhibitor of Drp1, over other dynamin family members, and reported to inhibit mitochondrial fission. A recent study has challenged the activity of mdivi-1 as an inhibitor of Drp1. This study raises serious issues regarding the interpretation of data addressing the effects of mdivi-1 as reflective of the inhibition of Drp1 and thus fission. This commentary considers the evidence for and against mdivi-1 as an inhibitor of Drp1 and presents the following considerations; (1) the activity of mdivi-1 toward Drp1 GTPase activity requires further biochemical investigation, (2) as there is a large body of literature using mdivi-1 in vitro with effects as predicted for inhibition of Drp1 and mitochondrial fission, reviewed herein, the evidence is in favor of mdivi-1's originally described bioactivity, and (3) until the issue is resolved, experimental interpretations for the effects of mdivi-1 on inhibition of fission in cell and tissue experiments warrants stringent positive controls directly addressing the effects of mdivi-1 on fission. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1260-1268, 2017.

Highly pathogenic avian influenza A virus H5N1 non-structural protein 1 is associated with apoptotic activation of the intrinsic mitochondrial pathway.

Outbreaks of avian influenza A (H5N1) virus infection have significant health and economic consequences. Non-structural protein 1 (NS1) is an essential virulence factor of the highly pathogenic H5N1 avian influenza virus and of the apoptosis associated with the pathogenesis of H5N1. Previous studies have revealed that the NS1 protein is able to induce apoptosis via an extrinsic pathway. However, it remains unclear whether the intrinsic pathway is also associated with this apoptosis. The present study used a clone of the NS1 gene from avian influenza A/Jiangsu/1/2007 and observed the localization of the NS1 protein and cytochrome c release from mitochondria and the change of mitochondrial membrane potential (MMP) in lung cancer cells. Cytotoxicity was detected using an MTT assay and the number of apoptotic cells was counted using a flow cytometer. Following the isolation of mitochondria, western blotting was performed to compare cytochrome c release from the mitochondria in cells before and after apoptosis. The change of MMP was detected using JC-1 staining. Furthermore, the results reveal that the majority of the NS1 protein was localized in the cell nucleus, and that it may induce apoptosis of human lung epithelial cells. The apoptosis occurred with marked cytochrome c release from mitochondria and a change of the MMP. This indicated that the NS1 protein may be associated with apoptosis induced by an intrinsic mitochondrial pathway.

Mitochondrial dysfunction is an acute response of articular chondrocytes to mechanical injury.

Direct therapeutic targeting of MT function in the early post-injury time frame may provide a strategy to block perpetuation of tissue damage and prevent the development of PTOA. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:739-750, 2018.

Mitochondrial expression and activity of P-glycoprotein under oxidative stress in outer blood-retinal barrier.

To investigate the role of oxidative stress in regulating the functional expression of P-glycoprotein (P-gp) in mitochondria of D407 cells. D407 cells were exposed to different ranges of concentrations of H2O2. The mitochondrial location of P-gp in the cells subjected to oxidative stress was detected by confocal analysis. Expression of P-gp in isolated mitochondria was assessed by Western blot. The pump activity of P-gp was evaluated by performing the efflux study on isolated mitochondria with Rhodamine 123 (Rho-123) alone and in the presence of P-gp inhibitor (Tariquidar) using flow cytometry analysis. The cells were pretreated with 10 mmol/L N-acetylcysteine (NAC) for 30min before exposing to H2O2, and analyzed the mitochondrial extracts by Western blot and flow cytometry. P-gp was co-localized in the mitochondria by confocal laser scanning microscopy, and it was also detected in the mitochondria of D407 cells using Western blot. Exposure to increasing concentrations of H2O2 led to gradually increased expression and location of P-gp in the mitochondria of cells. Rho-123 efflux assay showed higher uptake of Rho-123 on isolated mitochondria in the presence of Tariquidar both in normal and oxidative stress state. H2O2 up-regulated P-gp in D407 cells, which could be reversed by NAC treatment. H2O2 could up-regulate the functional expression of P-gp in mitochondria of D407 cells, while antioxidants might suppress oxidative-stress-induced over-expression of functional P-gp. It is indicative that limiting the mitochondrial P-gp transport in retinal pigment epithelium cells would be to improve the effect of mitochondria-targeted antioxidant therapy in age-related macular degeneration-like retinopathy.

Use of Extraction of Emblica Officinalis in Preparing Pharmaceutical Composition for Repairing Liver Damage

An use of an extraction of emblica officinalis in preparing pharmaceutical composition for repairing liver damage, wherein the pharmaceutical composition including the extraction of emblica officinalis improves the ability of performing the oxidative phosphorylation for the synthesis of the adenosine triphosphate (ATP) by mitochondria in the liver cells which are in contact with the free radicals. Thepharmaceutical composition including the extraction of emblica officinalis also reduces the proton leakage of the mitochondria, improves the maximal respiration of the mitochondria, improves the coupling efficiency of the adenosine triphosphate in the mitochondria, improves the spare respiratory capacity of the mitochondria, and reduces the amount of the free radical generated in the liver cells.

Effect of laser diode light irradiation on growth capability of human hair follicle dermal papilla cells.

Low level laser therapy is widely used to relieve pain and inflammation, and to restore cellular functions. The photons of light are absorbed by mitochondria in cells, leading to an increase in the production of adenosine triphosphate (ATP), nitric oxide release, blood flow, and reactive oxygen species (ROS). This study proposed the use of a laser diode array at 808 nm to stimulate the proliferation and to activate the functions of dermal papilla cells, which were an important part of the hair growth cycle. These cells were isolated from human hair follicles and were exposed to 808 nm light at various doses from 0.5, 1, 2.5, 4, and 6 J/cm2. The rate of cell proliferation and the gene expression profile of dermal papilla cells were investigated and compared with the control in which the cells did not received any light treatment. The growth curves of the dermal papilla cells were used to determine the specific growth rates. Higher specific growth rates were observed in the cells exposed to laser at doses higher than 0.5 J/cm2. The effect of the laser light treatment on several gene markers, specifically for dermal papilla cells, was evaluated using real-time polymerase chain reaction (qPCR). Our result shows that collagen type 1 (Col1), alkaline phosphatase (Alp), and versican (Vcan) did not increase when the cells were irradiated by the laser light. Interestingly, sex determining region y-box 2 (Sox2) gene was up-regulated when 0.5 J/cm2, and 1 J/cm2 light was used, while an increase in the level of fibroblast growth factor 7 (Fgf7) gene was observed with light irradiation at 0.5 J/cm2, 1 J/cm2, 2.5 J/cm2, and 4 J/cm2. Too high irradiation dose was shown to yield no effect on the gene expression of dermal papilla cells.