Reduced ATP Levels Research Articles

High Dietary Advanced Glycation End Products Impair Mitochondrial and Cognitive Function

Advanced glycation end products (AGEs) are an important risk factor for the development of cognitive decline in aging and late-onset neurodegenerative diseases including Alzheimer's disease. However, whether and how dietary AGEs exacerbate cognitive impairment and brain mitochondrial dysfunction in the aging process remains largely unknown. We investigated the direct effects of dietary AGEs on AGE adducts accumulation, mitochondrial function, and cognitive performance in mice. Mice were fed the AGE+ diet or AGE- diet. We examined levels of AGE adducts in serum and cerebral cortexes by immunodetection and immunohistochemistry, determined levels of reactive oxygen species by biochemical analysis, detected enzyme activity associated with mitochondrial respiratory chain complexes I & IV and ATP levels, and assessed learning and memory ability by Morris Water Maze and nesting behavior. Levels of AGE adducts (MG-H1 and CEL) were robustly increased in the serum and brain of AGE+ diet fed mice compared to the AGE- group. Furthermore, greatly elevated levels of reactive oxygen species, decreased activities of mitochondrial respiratory chain complexes I & IV, reduced ATP levels, and impaired learning and memory were evident in AGE+ diet fed mice compared to the AGE- group. These results indicate that dietary AGEs are important sources of AGE accumulation in vivo, resulting in mitochondrial dysfunction, impairment of energy metabolism, and subsequent cognitive impairment. Thus, reducing AGEs intake to lower accumulation of AGEs could hold therapeutic potential for the prevention and treatment of AGEs-induced mitochondrial dysfunction linked to cognitive decline.

2118-P: Regulation of Ataxia-Telangiectasia and Rad3-Related (ATR)–Dependent DNA Damage Response by Nitric Oxide in ß-Cells

Nitric oxide, generated in β-cells in response to cytokines, induces DNA damage that activates ataxia-telangiectasia mutated protein (ATM)-dependent DNA damage response (DDR). While the DDR does not contribute to DNA repair in β-cells, it does regulate β-cell apoptosis when the nitric oxide production falls below a threshold level. Indeed, nitric oxide at high micromolar levels, while inducing DNA damage, also inhibits DDR and protects β-cells from apoptosis. This ability of nitric oxide, a potent inhibitor of mitochondrial oxidation, to attenuate DDR activation correlates with a 10-fold reduction in ATP and the actions are selective to β-cells. Unlike most other cell types, β-cells cannot compensate for mitochondrial inhibition by increasing glycolysis, leading to a loss in ATP levels which correlates with DDR inhibition. We now show that nitric oxide also inhibits the activation of a second DDR transducer kinase, ATR. ATR is activated in response to single-strand DNA breaks and replication stress. We showed that low levels of nitric oxide activate ATR-dependent DDR signaling by inducing replication stress. However, much like with ATM, at higher concentrations nitric oxide and other inhibitors of mitochondrial respiration that reduce ATP levels by more than 10-fold, also inhibit ATR selectively in β-cells. Nitric oxide and these mitochondrial poisons do not inhibit ATR signaling in non-β-cells as they maintain ATP levels by increasing glycolysis under mitochondrial inhibition. Importantly, when non-β-cells are forced to utilize oxidative phosphorylation for ATP generation, nitric oxide and other mitochondrial inhibitors attenuate ATR signaling in a manner similar to β-cells. Together, these studies suggest that inhibition of the DDR by nitric oxide in β-cells is dependent on its ability to inhibit mitochondrial oxidative metabolism and deplete ATP, and suggest that the inhibition of DDR signaling may be a protective response to prevent β-cell apoptosis. Disclosure C. Yeo: None. B. Oleson: None. J.A. Corbett: None. J.K. Schnuck: None. Funding National Institutes of Health (DK052194, AI-44458)

The Recombinant Fragment of Human κ-Casein Induces Cell Death by Targeting the Proteins of Mitochondrial Import in Breast Cancer Cells.

Breast cancer is still one of the most common cancers for women. Specified therapeutics are indispensable for optimal treatment. In previous studies, it has been shown that RL2, the recombinant fragment of human κ-Casein, induces cell death in breast cancer cells. However, the molecular mechanisms of RL2-induced cell death remain largely unknown. In this study, mechanisms of RL2-induced cell death in breast cancer cells were systematically investigated. In particular, we demonstrate that RL2 induces loss of mitochondrial membrane potential and cellular ATP loss followed by cell death in breast cancer cells. The mass spectrometry-based screen for RL2 interaction partners identified mitochondrial import protein TOM70 as a target of RL2, which was subsequently validated. Further to this, we show that RL2 is targeted to mitochondria after internalization into the cells, where it can also be found in the dimeric form. The importance of TOM70 and RL2 interaction in RL2-induced reduction in ATP levels was validated by siRNA-induced downregulation of TOM70, resulting in the partial rescue of ATP production. Taken together, this study demonstrates that RL2–TOM70 interaction plays a key role in RL2-mediated cell death and targeting this pathway may provide new therapeutic options for treating breast cancer.

Di-2-picolylamine triggers caspase-independent apoptosis by inducing oxidative stress in human liver hepatocellular carcinoma cells.

Di-2-picolylamine (DPA) is an organic compound that has been shown to possess antioxidant properties when conjugated to form a metal complex. The basis of this study was to determine the effects of DPA on the proliferation and apoptosis of human hepatocellular carcinoma cells and elucidate the possible mechanisms. The methylthiazol tetrazolium assay served to measure cell viability and generated an IC50 of 1591 µM. Luminometry was used to investigate caspase activity and ATP concentration. It was observed that the decreased cell viability was associated with reduced ATP levels. Despite increased Bax and caspase 9 activity, cell death was caspase independent as indicated by the reduction in caspase 3/7 activity. This was associated with the downregulation poly(ADP-ribose) polymerase cleavage (Western blotting). However, the Hoescht assay depicted nuclear condensation and apoptotic body formation with elevated DPA levels suggesting DNA damage in HepG2 cells. DNA damage assessed by the comet assay confirmed an increased comet tail formation. The presence of oxidative stress was investigated by quantifying reactive species (malondialdehyde and nitrates concentration) and Western blotting to confirm the expression of antioxidant proteins. The DPA increased lipid peroxidation (RNS), a marker of oxidative stress, consequently causing cell death. The accompanying upregulation of stress-associated proteins superoxide dismutase (SOD2), nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and Hsp70 verifies oxidative stress.

Mitochondria Transfer Elicits Articular Cartilage Protection

Background & Aim Osteoarthritis (OA) is the most frequent joint disease worldwide with yet no known disease modifying drugs. In OA chondrocytes (OAC), the dysregulation of the metabolic biosensors that control mitochondrial biogenesis accounts for decreased numbers of mitochondria (MT) and reduced ATP levels in OA. Recently, our group evidenced that natural and artificial MT transfer (MitoT) from MSCs, convey potent metabolic and functional effects on target T cells, controlling organ damage and survival in an in-vivo model of inflammatory disease. In the present work, the effects of MitoT on the OAC phenotype is assessed in a coculture of MT-labelled MSCs with OAC. Methods, Results & Conclusion MitoT was evidenced as early as 4hrs and reached 97% after 24 hours. Cytochalasin B inhibition pointed to a tunneling Nanotubes (TNT)-dependent mechanism. The impact of MitoT, isolated from other effects of the MSC secretome was assessed in OACs incubated solely with MSC derived MT. FACS analysis revealed the internalization of exogenous MT with dose-dependent uptakes of MT equivalent to MSC:OAC ratios of 1:10, 1:5 and 1:1, with averages of 20.1%, 29.4% and 91.7% MitoTpos target OACs, respectively. Functional effect of the MitoT was assessed in readouts including (a) immunosuppression assays, where MitoTpos OAC induced only a slight change in PBMCs proliferation; (b) metabolic activity measured by WST-1 demonstrating an increase in the metabolic activity of MitoTpos OACs; (c) ROS-regulating autophagy gene expression levels showing a significant increase in SOD2 and Redd1mRNA levels in MitoTpos cells and (d) resistance to oxidative stress, assessed following H2O2 treatment, that revealed a protective effect of MitoT (15% reduction of dead cells). Furthermore, the intracellular fate of donated MT was evaluated with MT specific SNPs from the D-loop region. Using the amplification refractory mutation system (ARMS), a unique PCR product for MSC-derived MT was identified for at least 9 days in the target OACs. In conclusion, OAC were shown to be highly permissive for MT donation. The MitoT effect included an increase of metabolic activity and oxidative stress resistance that was correlated with an upregulation of SOD-2 expression. The biodistribution and efficacy of intraarticular injection of MT are currently being investigated in an OA mouse model. The MitoT induced reversal of the metabolic dysfunction typical of OACs might open new avenues to the understanding and treatment of OA.

OXIDATIVE STRESS AND MITOCHONDRIAL DYSFUNCTION

The process of cell damage resulting from the action of free radicals – reactive oxygen species (ROS) – is called oxidative stress. Most ROS are constantly formed in the cell – about 5 % of the oxygen consumed by tissues is converted into free radicals, but their level is normally so small that the cell inactivates them with the help of an antioxidant system. Different organs and tissues are exposed to different degrees of ROS and demonstrate different stability during the implementation of oxidative stress. The mechanisms of ROS formation by mitochondria under oxidative stress are still unclear. At the same time, it was found that mitochondrial dysfunction and the accumulation of mitochondrial mutations in tissues make a significant contribution to the aging process, as well as to the pathogenesis of a number of diseases characterized by neurodegeneration. Mutations lead to increased generation of free radicals, reduced ATP levels, and energy failure of cells. Coenzyme Q10 is a component of the mitochondrial respiratory chain. Violation of the biosynthesis of coenzyme Q10 can lead to a number of mitochondrial diseases. When coenzyme Q10 is deficient, sulfide metabolism plays a critical role. Sulfide metabolism in mammalian cells includes trans-sulfuration (biosynthetic) and hydrogen sulfide oxidation (H2S) (catabolic). Violation of H2S oxidation may contribute to oxidative stress in coenzyme Q deficiency or may play a synergistic role with oxidative stress in the pathogenesis of tissue specificity in coenzyme Q deficiency.

α-Mangostin protects against myocardial ischemia reperfusion injury by suppressing the activation of HIF-1α

Purpose: To investigate the cytoprotective effect of α-mangostin on myocardial tissues in ischemic rats, and the underlying mechanism.Methods: Histopathological changes in myocardial tissues were determined using inverted microscope. Protein expressions were measured by western blotting, while enzyme-linked immunosorbent assay (ELISA) was used to assay the expression levels of caspase-3, caspase-9 and caspase-8.Results: Treatment with α-mangostin (20 mg/kg) suppressed production of reactive oxygen species (ROS) and lipid peroxides in myocardial tissues of MI/R rats, and significantly alleviated MI/R injurymediated reduction in ATP levels in cardiac tissues (p < 0.05). α-Mangostin treatment of MI/R injury rats suppressed HIF-1α activation, and markedly elevated BNIP3 levels, relative to model group. Moreover, MI/R-induced cardiomyocyte apoptosis was significantly alleviated by α-mangostin treatment (p < 0.05). Treatment with α-mangostin also suppressed I/R-induced increases in caspase-8 and caspase-3 activation in myocardial tissues, improved Nrf-2 activation, and promoted HO-1 and GST levels in MI/R injury rats (p < 0.05).Conclusion: These results suggest that α-mangostin protects rat cardiac tissues from MI/R-induced oxidative damage via reduction of HIF-1α expression, inhibition of ROS generation and suppression of apoptosis. Therefore, α-mangostin may be of therapeutic importance for the management of myocardial ischemia in humans.
 Keywords: α-Mangostin, Hypoxia, Inflammation, Nrf-2, Oxidative stress, Reperfusion

Pseudomonas putida KT2440 is naturally endowed to withstand industrial-scale stress conditions.

Summary Pseudomonas putida is recognized as a very promising strain for industrial application due to its high redox capacity and frequently observed tolerance towards organic solvents. In this research, we studied the metabolic and transcriptional response of P. putida KT2440 exposed to large‐scale heterogeneous mixing conditions in the form of repeated glucose shortage. Cellular responses were mimicked in an experimental setup comprising a stirred tank reactor and a connected plug flow reactor. We deciphered that a stringent response‐like transcriptional regulation programme is frequently induced, which seems to be linked to the intracellular pool of 3‐hydroxyalkanoates (3‐HA) that are known to serve as precursors for polyhydroxyalkanoates (PHA). To be precise, P. putida is endowed with a survival strategy likely to access cellular PHA, amino acids and glycogen in few seconds under glucose starvation to obtain ATP from respiration, thereby replenishing the reduced ATP levels and the adenylate energy charge. Notably, cells only need 0.4% of glucose uptake to build those 3‐HA‐based energy buffers. Concomitantly, genes that are related to amino acid catabolism and β‐oxidation are upregulated during the transient absence of glucose. Furthermore, we provide a detailed list of transcriptional short‐ and long‐term responses that increase the cellular maintenance by about 17% under the industrial‐like conditions tested.

Novel benzoate-lipophilic cations selectively induce cell death in human colorectal cancer cell lines

IntroductionColorectal cancer (CRC) is a critical health issue worldwide. The high rate of liver and lung metastasis associated with CRC creates a significant barrier to effective and efficient therapy. Tumour cells, including CRC cells, have metabolic alterations, such as high levels of glycolytic activity, increased cell proliferation and invasiveness, and chemo- and radio-resistance. However, the abnormally elevated mitochondrial transmembrane potential of these cells also provides an opportunity to develop drugs that selectively target the mitochondrial functions of tumour cells. MethodsIn this work, we used a new batch of benzoic acid esters with cytotoxic activities attached to the triphenylphosphonium group as a vehicle to target tumour mitochondria and improve their activity. We evaluated the cytotoxicity, selectivity, and mechanism of action of these derivatives, including the effects on energy stress-induced apoptosis and metabolic behaviour in the human CRC cell lines HCT-15 and COLO-205. ResultsThe benzoic acid derivatives selectively targeted the tumour cells with high potency and efficacy. The derivatives induced the uncoupling of the oxidative phosphorylation system, decreased the transmembrane potential, and reduced ATP levels while increasing AMPK activation, thereby triggering tumour cell apoptosis in both tumour cell lines tested. ConclusionThe benzoic acid derivatives studied here are promising candidates for assessing in vivo models of CRC, despite the diverse metabolic characteristics of these tumour cells.

Nrf2 activation through the PI3K/GSK-3 axis protects neuronal cells from A\u03b2-mediated oxidative and metabolic damage

BackgroundMounting evidence points to a crucial role of amyloid-β (Aβ) in the pathophysiology of Alzheimer’s disease (AD), a disorder in which brain glucose hypometabolism, downregulation of central elements of phosphorylation pathways, reduced ATP levels, and enhanced oxidative damage coexist, and sometimes precede, synaptic alterations and clinical manifestations. Since the brain has limited energy storage capacity, mitochondria play essential roles in maintaining the high levels of energy demand, but, as major consumers of oxygen, these organelles are also the most important generators of reactive oxygen species (ROS). Thus, it is not surprising that mitochondrial dysfunction is tightly linked to synaptic loss and AD pathophysiology. In spite of their relevance, the mechanistic links among ROS homeostasis, metabolic alterations, and cell bioenergetics, particularly in relation to Aβ, still remain elusive.MethodsWe have used classic biochemical and immunocytochemical approaches together with the evaluation of real-time changes in global energy metabolism in a Seahorse Metabolic Analyzer to provide insights into the detrimental role of oligAβ in SH-SY5Y and primary neurons testing their pharmacologic protection by small molecules.ResultsOur findings indicate that oligomeric Aβ induces a dramatic increase in ROS production and severely affects neuronal metabolism and bioenergetics. Assessment of global energy metabolism in real time demonstrated Aβ-mediated reduction in oxygen consumption affecting basal and maximal respiration and causing decreased ATP production. Pharmacologic targeting of Aβ-challenged neurons with a set of small molecules of known antioxidant and cytoprotective activity prevented the metabolic/bioenergetic changes induced by the peptide, fully restoring mitochondrial function while inducing an antioxidant response that counterbalanced the ROS production. Search for a mechanistic link among the protective small molecules tested identified the transcription factor Nrf2—compromised by age and downregulated in AD and transgenic models—as their main target and the PI3K/GSK-3 axis as the central pathway through which the compounds elicit their Aβ protective action.ConclusionsOur study provides insights into the complex molecular mechanisms triggered by oligAβ which profoundly affect mitochondrial performance and argues for the inclusion of small molecules targeting the PI3K/GSK-3 axis and Nrf2-mediated pathways as part of the current or future combinatorial therapies.

Decreasing microbially influenced metal corrosion using free nitrous acid in a simulated water injection system

Microbially influenced corrosion (MIC) is the main cause of metal corrosion in anoxic environments. Biocides are often dosed to the corrosive media to inhibit and kill the microbes which cause MIC. In this study, intermittent dosages of free nitrous acid (FNA), which was previously found to be a biocide, were applied to a simulated water injection system containing carbon steel coupons with mature biofilm, to study the effect of FNA on mitigation of metal corrosion. In each treatment, 0.49 mg-N/L FNA was dosed using 200 mg-N/L nitrite at pH 6 for 24 h. The corrosion properties were monitored by open circuit potential (OCP), electrochemical impedance spectroscopy (EIS), linear polarization resistance (LPR), 3D optical profiling, and direct weight measurement. The biofilm viability was monitored by measuring cellular ATP level. The general corrosion rate (calculated by weight-loss measurement) was decreased by up to 31%, which was supported by LPR tests and reduced ATP levels of the corrosion-inducing biofilm. The 3D optical profiling results showed that FNA decreased the average pitting corrosion rate by 59%, with 2 intermittent treatments and 82-day interval over 304 days. Intermittent dosing of FNA has strong potential to be an effective and efficient strategy for controlling MIC in oil recovery infrastructure.

The mgtCBR mRNA Leader Secures Growth of Salmonella in Both Host and Non-host Environments.

Upon intracellular cues, bacterial mRNA leaders often form secondary structures that determine expression of a downstream protein-coding region(s), thereby providing bacteria with a mechanism to control the amounts of necessary proteins in the right locales. Here we describe a polycistronic mRNA leader that secures bacterial growth by preventing dysregulated expression of the protein-coding regions. In Salmonella, the mgtCBR mRNA encodes the virulence protein MgtC and the Mg2+ transporter MgtB. A mutant designed to produce leaderless mgtCBR mRNA induced MgtC and MgtB in conditions that promote mgtC transcription. The dysregulated expression of MgtC and MgtB impaired bacterial growth under all such non-host environments. While MgtC, but not MgtB, normally reduces ATP levels in a process requiring the F1F0 ATP synthase, dysregulated MgtC and MgtB reduced ATP levels independently of the F1F0 ATP synthase, which correlated with the mutant’s growth defect. The mutant showed dysregulated MgtC expression and attenuated survival inside macrophages. While MgtB normally does not affect the phenotype, MgtB impaired intramacrophage survival of the mutant in the presence of MgtC. We provide an example showing that a polycistronic mRNA leader prevents the dysregulated function of protein-coding regions to allow bacteria to proliferate across complex niches.

Artemesia Annua L. Nanoparticles Destabilize Membrane Integrity and Induce Apoptosis in a Caspase Mediated Pathway in MDA-MB-231 Cells

Breast cancer exhibits diagnostic and therapeutic challenges due to tumour heterogenicity and poor efficacy of anti cancer drugs at the metastatic sites. Recently, nanomedicines provide a potential benefit to breast cancer treatment by their increased bioavailability, selective targeting, increased efficacy and lower toxicity. This work aimed to evaluate the cytotoxic potential of A. annua silver nanoparticles (AGNPs) on human triple-negative breast cancer cell lines. The mechanism by which A.annua AGNPs exhibit growth inhibition was analysed by using the status of LDH, Mitochondria Membrane potential [MMP], and ATP levels. Finally, the ability of A. annua AGNPs to mediate cell death was studied by protein expressions of Bax, BCL-2, Caspase 3and Caspase 9. We observed that A. annua AGNPs show a remarkable cytotoxic effect by increasing levels of LDH with loss of MMP and reduced ATP levels. Here in this study, protein expression of Bax, BCL-2 ratio, Cyt C and Caspase 9 protein expression level favors apoptotic induction during AGNPs treatment. Henceforth we prove that A.annua AGNPs can be useful as alleviative regiment against breast cancer and can be explored for other cancers and its related pathologies.

BRCA1 Deficiency Upregulates NNMT, Which Reprograms Metabolism and Sensitizes Ovarian Cancer Cells to Mitochondrial Metabolic Targeting Agents.

BRCA1 plays a key role in homologous recombination (HR) DNA repair. Accordingly, changes that downregulate BRCA1, including BRCA1 mutations and reduced BRCA1 transcription, due to promoter hypermethylation or loss of the BRCA1 transcriptional regulator CDK12, disrupt HR in multiple cancers. In addition, BRCA1 has also been implicated in the regulation of metabolism. Here, we show that reducing BRCA1 expression, either by CDK12 or BRCA1 depletion, led to metabolic reprogramming of ovarian cancer cells, causing decreased mitochondrial respiration and reduced ATP levels. BRCA1 depletion drove this reprogramming by upregulating nicotinamide N-methyltransferase (NNMT). Notably, the metabolic alterations caused by BRCA1 depletion and NNMT upregulation sensitized ovarian cancer cells to agents that inhibit mitochondrial metabolism (VLX600 and tigecycline) and to agents that inhibit glucose import (WZB117). These observations suggest that inhibition of energy metabolism may be a potential strategy to selectively target BRCA1-deficient high-grade serous ovarian cancer, which is characterized by frequent BRCA1 loss and NNMT overexpression. SIGNIFICANCE: Loss of BRCA1 reprograms metabolism, creating a therapeutically targetable vulnerability in ovarian cancer.

Oxidative Phosphorylation Regulated By PGC-1α Promotes Multiple Myeloma Progression

Introduction: With the deeper understanding of metabolic reprograming, more and more studies highlight that many tumors heavily rely on mitochondrial oxidative phosphorylation (OXPHOS) for bioenergetics and biosynthesis, and targeting OXPHOS appears to be a promising approach for cancer therapy. Despite multiple metabolism alterations occur in multiple myeloma (MM), the role of OXPHOS in MM is rarely focused on. Many metabolic pathways are orchestrated by certain master co-regulators through transcriptional programs. The expression of OXPHOS gene set was reported to be regulated by peroxisome proliferator activated receptor gamma coactivator 1α (PGC-1α) in some other tissues, which serves as a transcriptional coactivator. Our previous studies demonstrated that PGC-1α was overexpression in MM cell line RPMI-8226 and knockdown of PGC-1α inhibited proliferation. Herein, we aimed to investigate the significance of OXPHOS in MM and clarify whether PGC-1α is the core regulator of OXPHOS in MM. Methods: Transcriptome data of GSE6477, GSE13591, GSE47552 were downloaded from the Gene Expression Omnibus (GEO) database and then removed batch effects. Gene set variation analysis (GSVA) were used to identify pathways enriched in MM. Differential expression analysis was performed and the PGC-1α expression value was extracted. Gene set enrichment analysis (GSEA) was adopted to explore biological functions involved by PGC-1α. RT-PCR was conducted to reflect the influence of PGC-1α inhibitor on OXPHOS gene set. Mitochondrial morphology was exhibited by transmission electron microscopy analysis. Cell counting kit-8 assay was employed to determine cell viability and flow cytometry analysis was used to assess apoptosis. Results: Integrated bioinformatics analysis revealed that OXPHOS pathway was significantly upregulated in MM than in normal donors (Figure A), and aberrantly overexpression of OXPHOS gene set was confirmed in our MM samples at mRNA level (Figure B-C). Moreover, we also demonstrated that aberrantly overexpression of OXPHOS genes in MM were generally associated with poorer survival (Figure D-F), indicating that OXPHOS appears as a potential oncogenic pathway for MM progression. Meanwhile, bioinformatics results showed that PGC-1α was upregulated in 247 MM patients from GEO database (Figure G). In addition, our results indicated that the overexpression of PGC-1α prevailed among 5 MM cell lines with different inherited backgrounds and our several MM samples (Figure H-I). GSEA analysis showed that gene sets of OXPHOS, TCA cycle, respiratory electron transport, ATP synthesis, were all significantly enriched in MM patients with high PGC-1α expression (Figure J-L). The inhibition of PGC-1α have exerted significant inhibition effect on the transcription of OXPHOS gene set in MM. In accordance with that OXPHOS is a major source of ATP, PGC-1α inhibitor resulted in the reduction of ATP levels of MM cells. Meanwhile, treatment of PGC-1α inhibitor triggered shriveled mitochondria with decreased volume, disorganized cristae, and increased electron density of matrix, indicative of impaired mitochondria OXPHOS. In vitro experiments suggested that PGC-1α inhibitor robustly inhibited proliferation of MM cells in a time and dose-dependent manner, with little effect on normal hematopoietic cells. Besides, PGC-1α inhibitor induced MM cells apoptosis significantly. Conclusions: Our investigations reported for the first time that OXPHOS regulated by PGC-1α may serve as a potential mechanism for MM progression. Besides, this study may provide new strategies for the treatment of MM from the perspective of OXPHOS and its core regulator. Figure Disclosures No relevant conflicts of interest to declare.

Toxicity assessment of the herbicide acetochlor in the human liver carcinoma (HepG2) cell line.

Acetochlor is a high-volume herbicide used on a global scale and toxicity assessments are needed to define its potential for adverse effects in wildlife and humans. This study was conducted to determine the effects of acetochlor on human liver carcinoma cells (HepG2), a cell model widely used to assess the potential for chemical hepatotoxicity. Experiments were conducted at concentrations ranging 0-800 μM acetochlor over a 12 to 48h period to quantify underlying mechanisms of toxicity. Our data indicate that acetochlor suppressed HepG2 cell proliferation in both a concentration- and time-dependent manner. Acetochlor induced reactive oxygen species (ROS) generation more than 700% with exposure to 400 μM acetochlor, and acetochlor decreased the activities and levels of anti-oxidant responses (superoxide dismutase, glutathione) following exposure to 100 μM, 200 μM and 400 μM acetochlor. Acetochlor also (1) induced HepG2 cell damage through apoptotic-signaling pathways; (2) enhanced intracellular free Ca2+ concentration (>400%); (3) decreased mitochondrial transmembrane potential (∼77%), and reduced ATP levels (∼65%) following exposure to 400 μM acetochlor compared to untreated cells. Notably, cell cycle progression was blocked at G0/G1 phase in HepG2 cells when treated for 24 h with 400 μM acetochlor. Taken together, acetochlor induced significant cytotoxicity toward HepG2 cells, and the underlying toxicity mechanisms appear to be related to ROS generation, mitochondrial dysfunction and disruption in the cell cycle regulation. These data contribute to toxicity assessments for acetochlor, a high-use herbicide, to quantify risk to wildlife and human health.

Reductions in COQ2 Expression Relate to Reduced ATP Levels in Multiple System Atrophy Brain.

Multiple system atrophy (MSA) is a progressive neurodegenerative disease clinically characterized by parkinsonism and cerebellar ataxia, and pathologically by oligodendrocyte α-synuclein inclusions. Genetic variants of COQ2 are associated with an increased risk for MSA in certain populations. Also, deficits in the level of coenzyme Q10 and its biosynthetic enzymes are associated with MSA. Here, we measured ATP levels and expression of biosynthetic enzymes for coenzyme Q10, including COQ2, in multiple regions of MSA and control brains. We found a reduction in ATP levels in disease-affected regions of MSA brain that associated with reduced expression of COQ2 and COQ7, supporting the concept that abnormalities in the biosynthesis of coenzyme Q10 play an important role in the pathogenesis of MSA.

Mitochondrial damage by \u03b1-synuclein causes cell death in human dopaminergic neurons

Evolving concepts on Parkinson’s disease (PD) pathology suggest that α-synuclein (aSYN) promote dopaminergic neuron dysfunction and death through accumulating in the mitochondria. However, the consequence of mitochondrial aSYN localisation on mitochondrial structure and bioenergetic functions in neuronal cells are poorly understood. Therefore, we investigated deleterious effects of mitochondria-targeted aSYN in differentiated human dopaminergic neurons in comparison with wild-type (WT) aSYN overexpression and corresponding EGFP (enhanced green fluorescent protein)-expressing controls. Mitochondria-targeted aSYN enhanced mitochondrial reactive oxygen species (ROS) formation, reduced ATP levels and showed severely disrupted structure and function of the dendritic neural network, preceding neuronal death. Transmission electron microscopy illustrated distorted cristae and many fragmented mitochondria in response to WT-aSYN overexpression, and a complete loss of cristae structure and massively swollen mitochondria in neurons expressing mitochondria-targeted aSYN. Further, the analysis of mitochondrial bioenergetics in differentiated dopaminergic neurons, expressing WT or mitochondria-targeted aSYN, elicited a pronounced impairment of mitochondrial respiration. In a pharmacological compound screening, we found that the pan-caspase inhibitors QVD and zVAD-FMK, and a specific caspase-1 inhibitor significantly prevented aSYN-induced cell death. In addition, the caspase inhibitor QVD preserved mitochondrial function and neuronal network activity in the human dopaminergic neurons overexpressing aSYN. Overall, our findings indicated therapeutic effects by caspase-1 inhibition despite aSYN-mediated alterations in mitochondrial morphology and function.

Expression of Human Mutant Huntingtin Protein in Drosophila Hemocytes Impairs Immune Responses.

The pathogenic effect of mutant HTT (mHTT) which causes Huntington disease (HD) are not restricted to nervous system. Such phenotypes include aberrant immune responses observed in the HD models. However, it is still unclear how this immune dysregulation influences the innate immune response against pathogenic infection. In the present study, we used transgenic Drosophila melanogaster expressing mutant HTT protein (mHTT) with hemocyte-specific drivers and examined the immune responses and hemocyte function. We found that mHTT expression in the hemocytes did not affect fly viability, but the numbers of circulating hemocytes were significantly decreased. Consequently, we observed that the expression of mHTT in the hemocytes compromised the immune responses including clot formation and encapsulation which lead to the increased susceptibility to entomopathogenic nematode and parasitoid wasp infections. In addition, mHTT expression in Drosophila macrophage-like S2 cells in vitro reduced ATP levels, phagocytic activity and the induction of antimicrobial peptides. Further effects observed in mHTT-expressing cells included the altered production of cytokines and activation of JAK/STAT signaling. The present study shows that the expression of mHTT in Drosophila hemocytes causes deficient cellular and humoral immune responses against invading pathogens. Our findings provide the insight into the pathogenic effects of mHTT in the immune cells.

Silica nanoparticles exacerbates reproductive toxicity development in high-fat diet-treated Wistar rats

To demonstrate the combined adverse effect and the mechanism of silica nanoparticles (SiNPs) with 57.66 ± 7.30 nm average diameter and high-fat diet (HFD) on Wistar rats, 60 male Wistar rats were randomly divided into six groups (n = 10): Control group, SiNPs group, HFD group, 2 mg kg−1 SiNPs + HFD group, 5 mg kg−1 SiNPs + HFD group and 10 mg kg−1 SiNPs + HFD group. HFD was administrated for 2 weeks for the rats in advance and SiNPs were supplied every 3 d for 48 d subsequently. The present study illustrated that both HFD and SiNPs could decrease sperm concentration, mobility rates, increase abnormality rates, damage testicular structure, reduce spermatogonium numbers and spermatoblast numbers, reduce ATP levels, and affect expression of regulatory factors for meiosis in testis. HFD and SiNPs further damaged the sperm and lowered the ATP level and expression of factors associated with meiotic signaling pathway compared with the HFD without SiNPs in testicular tissue of Wistar rats. These results suggested that SiNPs significantly promoted reproductive toxicity induced by HFD in Wistar rats, which provides novel experimental evidence and an explanation for magnified reproductive toxicity triggered by SiNPs in HFD rats.