Modulation Of Mitochondrial Metabolism Research Articles

Mitochondrial ribosomal protein L12 potentiates hepatocellular carcinoma by regulating mitochondrial biogenesis and metabolic reprogramming

BackgroundMitochondrial dysfunction and metabolic reprogramming are key features of hepatocellular carcinoma (HCC). Despite its significance, the precise underlying mechanism behind these processes has not been fully elucidated. The latest investigations, along with our previous discoveries, have substantiated the significant role of mitochondrial ribosomal protein L12 (MRPL12), a newly identified gene involved in mitochondrial transcription regulation, in the modulation of mitochondrial metabolism. Nevertheless, the role of MRPL12 in tumorigenesis has yet to be investigated. MethodsThe expression of MRPL12 in HCC was assessed using an online database. Western blot, quantitative real-time polymerase chain reaction (qRT-PCR), and immunohistochemistry (IHC) were employed to determine the expression of MRPL12 in HCC tissues, patient-derived organoid (PDO), and cell lines. The correlation between MRPL12 expression and clinicopathological features, as well as prognosis, was examined using tissue microarray analysis. An in vivo subcutaneous tumor xenograft model, gene knockdown or overexpression assay, chromatin immunoprecipitation (ChIP) assay, Seahorse XF96 assay, and cell function assay were employed to investigate the biological function and potential molecular mechanism of MRPL12 in HCC. ResultsA significant upregulation of MRPL12 was observed in HCC cells, PDO and patient tissues, which correlated with advanced tumor stage, higher grade and poor prognosis. MRPL12 overexpression promoted cell proliferation, migration, and invasion in vitro, as well as tumorigenicity in vivo, whereas MRPL12 knockdown showed the opposite effect. MRPL12 knockdown also inhibited the capacity of organoids proliferation capacity. Furthermore, MRPL12 was found to be crucial for maintaining mitochondrial homeostasis. Both gain and loss-of-function experiments targeting MRPL12 in HCC cells altered oxidative phosphorylation (OXPHOS) and mitochondrial DNA content. Notably, suppression of OXPHOS effectively mitigates the tumor-promoting effect attributed to MRPL12 overexpression, implying the involvement of MRPL12 in HCC through the modulation of mitochondrial metabolism. Besides, Yin Yang 1 (YY1) was identified as a transcription factor responsible for regulating MRPL12, while the PI3K/mTOR pathway was found to act as an upstream regulator of YY1. MRPL12 knockdown attenuated the YY1 overexpression or PI3K/mTOR activation-induced malignant phenotype in HCC cells. ConclusionOur findings provide compelling evidence that MRPL12 is implicated in driving the malignant phenotype of HCC via regulating mitochondrial metabolism. Moreover, the aberrant expression of MRPL12 in HCC is mediated by the upstream PI3K/mTOR/YY1 pathway. These results highlight the potential of targeting MRPL12 as a promising therapeutic strategy for the treatment of HCC.

Abstract P3193: Cardiac Glutamine Metabolism As NAD+ Generating Pathway During Chronic Hypoxia

Cardiovascular disease morality relies in the inability of the adult mammalian heart to regenerate damaged tissue due to the inability of the adult cardiomyocytes to proliferate. This lack of proliferation in the adult heart is due to an increased DNA damage and activation of the DNA Damage response upon an increase of mitochondrial ROS generation. We have previously shown that modulation of mitochondrial metabolism towards glucose oxidation at expenses of fatty acid utilization can induce adult cardiomyocyte proliferation and support cardiac regeneration. Moreover, by modulating the oxygen levels and, thus, cardiomyocyte metabolism, by chronic hypoxia exposure, we were also able to induce cardiomyocyte proliferation and cardiac regeneration in adult mice. Remarkably, our metabolomic studies of the hypoxic hearts did not show increase of lactate production, a hallmark of hypoxic metabolism in order to regenerate NAD+ required for glycolysis and ATP production under low oxygen conditions. Interestingly, glutamine metabolism is one of the most compensatory pathways in these conditions. Glutamine can be metabolized to its conversion into glutamate, which can enter in the Krebs’ Cycle, but also feed the malate-aspartate shuttle. This shuttle has the ability to generate cytoplasmic NAD+. Here, we hypothesize that, during chronic hypoxia exposure, the heart uses glutamine as a fuel to regenerate cytoplasmic NAD+ as an alternative to lactate production. By means of carbon labeling and metabolic flux studies in vivo, we have proven that glutamine contribution to the malate-aspartate shuttle is enhanced during hypoxia in the heart. Moreover, we have determined that glutamine supplementation during hypoxia in cell culture increases cytoplasmid NAD+ levels, confirming the contribution of glutamine catabolism to NAD/NADH balance during hypoxia. Overall, we show that glutamine metabolism is an important adaptation to hypoxic metabolism in the heart and describe a new mechanism of metabolic rewiring under hypoxia beyond the classic lactate production upon glycolysis upregulation.

Pharmacological Modulation of Energy and Metabolic Pathways Protects Hearing in the Fus1/Tusc2 Knockout Model of Mitochondrial Dysfunction and Oxidative Stress.

Tightly regulated and robust mitochondrial activities are critical for normal hearing. Previously, we demonstrated that Fus1/Tusc2 KO mice with mitochondrial dysfunction exhibit premature hearing loss. Molecular analysis of the cochlea revealed hyperactivation of the mTOR pathway, oxidative stress, and altered mitochondrial morphology and quantity, suggesting compromised energy sensing and production. Here, we investigated whether the pharmacological modulation of metabolic pathways using rapamycin (RAPA) or 2-deoxy-D-glucose (2-DG) supplementation can protect against hearing loss in female Fus1 KO mice. Additionally, we aimed to identify mitochondria- and Fus1/Tusc2-dependent molecular pathways and processes critical for hearing. We found that inhibiting mTOR or activating alternative mitochondrial energetic pathways to glycolysis protected hearing in the mice. Comparative gene expression analysis revealed the dysregulation of critical biological processes in the KO cochlea, including mitochondrial metabolism, neural and immune responses, and the cochlear hypothalamic-pituitary-adrenal axis signaling system. RAPA and 2-DG mostly normalized these processes, although some genes showed a drug-specific response or no response at all. Interestingly, both drugs resulted in a pronounced upregulation of critical hearing-related genes not altered in the non-treated KO cochlea, including cytoskeletal and motor proteins and calcium-linked transporters and voltage-gated channels. These findings suggest that the pharmacological modulation of mitochondrial metabolism and bioenergetics may restore and activate processes critical for hearing, thereby protecting against hearing loss.

Sirtuins and redox signaling interplay in neurogenesis, neurodegenerative diseases, and neural cell reprogramming.

Since the discovery of Neural Stem Cells (NSCs) there are still mechanism to be clarified, such as the role of mitochondrial metabolism in the regulation of endogenous adult neurogenesis and its implication in neurodegeneration. Although stem cells require glycolysis to maintain their stemness, they can perform oxidative phosphorylation and it is becoming more and more evident that mitochondria are central players, not only for ATP production but also for neuronal differentiation's steps regulation, through their ability to handle cellular redox state, intracellular signaling, epigenetic state of the cell, as well as the gut microbiota-brain axis, upon dietary influences. In this scenario, the 8-oxoguanine DNA glycosylase (OGG1) repair system would link mitochondrial DNA integrity to the modulation of neural differentiation. On the other side, there is an increasing interest in NSCs generation, from induced pluripotent stem cells, as a clinical model for neurodegenerative diseases (NDs), although this methodology still presents several drawbacks, mainly related to the reprogramming process. Indeed, high levels of reactive oxygen species (ROS), associated with telomere shortening, genomic instability, and defective mitochondrial dynamics, lead to pluripotency limitation and reprogramming efficiency's reduction. Moreover, while a physiological or moderate ROS increase serves as a signaling mechanism, to activate differentiation and suppress self-renewal, excessive oxidative stress is a common feature of NDs and aging. This ROS-dependent regulatory effect might be modulated by newly identified ROS suppressors, including the NAD+-dependent deacetylase enzymes family called Sirtuins (SIRTs). Recently, the importance of subcellular localization of NAD synthesis has been coupled to different roles for NAD in chromatin stability, DNA repair, circadian rhythms, and longevity. SIRTs have been described as involved in the control of both telomere's chromatin state and expression of nuclear gene involved in the regulation of mitochondrial gene expression, as well as in several NDs and aging. SIRTs are ubiquitously expressed in the mammalian brain, where they play important roles. In this review we summarize the current knowledge on how SIRTs-dependent modulation of mitochondrial metabolism could impact on neurogenesis and neurodegeneration, focusing mainly on ROS function and their role in SIRTs-mediated cell reprogramming and telomere protection.

DUT enhances drug resistance to proteasome inhibitors via promoting mitochondrial function in multiple myeloma.

Acquired chemoresistance to proteasome inhibitors (PIs), such as bortezomib (BTZ), becomes an intractable obstacle in the management of multiple myeloma (MM) in the clinic, but the underlying mechanisms are still not well elucidated. In the current study, we established bortezomib-resistant (BR) myeloma cells and performed stable isotope labeling by amino acids in cell culture (SILAC) assay to screen profiled protein expression. The level of deoxyuridine triphosphatase (DUT), an important enzyme of nucleotide metabolism, increased in the BR MM cells. Retrospective analysis indicated patients with higher DUT expression had poorer responses to PI-based treatment and clinical outcomes. DUT knockdown by RNAi effectively minimized BTZ resistance in MM cells. Moreover, DUT knockdown was accompanied with the downregulation of proliferating cell nuclear antigen (PCNA), contributing to decelerating cell growth, as well as augmented apoptosis due to bortezomib treatment. In contrast, DUT overexpression in parental MM.1S and LP-1 cells enhanced BTZ resistance. Furthermore, acquired resistance to BTZ could trigger the modulation of mitochondrial metabolism and function, as evidenced by elevated expression of genes associated with mitochondrial metabolism, as well as altered oxygen consumption rate and adenosine triphosphate (ATP) production in BR MM cells. DUT inhibition partially attenuated mitochondrial modulation, and instead favored an early impairment of mitochondrial integrity upon BTZ exposure so as to restrict MM progression and overcome drug resistance to BTZ treatment both in vitro and in vivo. In conclusion, we unveiled previously unrecognized effects of DUT on acquired drug resistance of MM, thus manipulating DUT may be efficacious for sensitizing MM cells to PIs.

A Proof-of-Concept Inhibitor of Endothelial Lipase Suppresses Triple-Negative Breast Cancer Cells by Hijacking the Mitochondrial Function.

Simple SummaryEndothelial lipase (EL/LIPG) is a key regulator of tumor cell metabolism. In triple-negative breast cancer (TNBC) cells, we find that the expression of LIPG is associated with long non-coding RNA DANCR and positively correlates with gene signatures of mitochondrial metabolism-oxidative phosphorylation (OXPHOS). DANCR binds to LIPG, which enables tumor cells to maintain the expression. Importantly, LIPG knockdown inhibits OXPHOS and TNBC tumor formation. Finally, our study identifies a natural compound, the LIPG inhibitor cynaroside, which provides a new therapeutic strategy against TNBC.Triple-negative breast cancer (TNBC) cells reprogram their metabolism to provide metabolic flexibility for tumor cell growth and survival in the tumor microenvironment. While our previous findings indicated that endothelial lipase (EL/LIPG) is a hallmark of TNBC, the precise mechanism through which LIPG instigates TNBC metabolism remains undefined. Here, we report that the expression of LIPG is associated with long non-coding RNA DANCR and positively correlates with gene signatures of mitochondrial metabolism-oxidative phosphorylation (OXPHOS). DANCR binds to LIPG, enabling tumor cells to maintain LIPG protein stability and OXPHOS. As one mechanism of LIPG in the regulation of tumor cell oxidative metabolism, LIPG mediates histone deacetylase 6 (HDAC6) and histone acetylation, which contribute to changes in IL-6 and fatty acid synthesis gene expression. Finally, aided by a relaxed docking approach, we discovered a new LIPG inhibitor, cynaroside, that effectively suppressed the enzyme activity and DANCR in TNBC cells. Treatment with cynaroside inhibited the OXPHOS phenotype of TNBC cells, which severely impaired tumor formation. Taken together, our study provides mechanistic insights into the LIPG modulation of mitochondrial metabolism in TNBC and a proof-of-concept that targeting LIPG is a promising new therapeutic strategy for the treatment of TNBC.

Quantitative Metabolomics of Tissue, Perfusate, and Bile from Rat Livers Subjected to Normothermic Machine Perfusion.

Machine perfusion (MP) allows the maintenance of liver cells in a metabolically active state ex vivo and can potentially revert metabolic perturbations caused by donor warm ischemia, procurement, and static cold storage (SCS). The present preclinical research investigated the metabolic outcome of the MP procedure by analyzing rat liver tissue, bile, and perfusate samples by means of high-field (600 MHz) nuclear magnetic resonance (NMR) spectroscopy. An established rat model of normothermic MP (NMP) was used. Experiments were carried out with the addition of an oxygen carrier (OxC) to the perfusion fluid (OxC-NMP, n = 5) or without (h-NMP, n = 5). Bile and perfusate samples were collected throughout the procedure, while biopsies were only taken at the end of NMP. Two additional groups were: (1) Native, in which tissue or bile specimens were collected from rats in resting conditions; and (2) SCS, in which biopsies were taken from cold-stored livers. Generally, NMP groups showed a distinctive metabolomic signature in all the analyzed biological matrices. In particular, many of the differentially expressed metabolites were involved in mitochondrial biochemical pathways. Succinate, acetate, 3-hydroxybutyrate, creatine, and O-phosphocholine were deeply modulated in ex vivo perfused livers compared to both the Native and SCS groups. These novel results demonstrate a broad modulation of mitochondrial metabolism during NMP that exceeds energy production and redox balance maintenance.

Mitochondrial bioenergetic pathways in blood leukocyte transcriptome decrease after intensive weight loss but are rescued following weight regain in female physique athletes.

Prolonged periods of energy deficit leading to weight loss induce metabolic adaptations resulting in reduced energy expenditure, but the mechanisms for energy conservation are incompletely understood. We examined 42 healthy athletic females (age 27.5±4.0years, body mass index 23.4±1.7kg/m2 ) who volunteered into either a group dieting for physique competition (n=25) or a control group (n=17). The diet group substantially reduced their energy intake and moderately increased exercise levels to induce loss of fat mass that was regained during a voluntary weight regain period. The control group maintained their typical lifestyle habits and body mass as instructed. From the diet group, fasting blood samples were drawn at baseline (PRE), after 4- to 5-month weight loss (PRE-MID), and after 4- to 5-month weight regain (MID-POST) as well as from the control group at similar intervals. Blood was analyzed to determine leukocyte transcriptome by RNA-Sequencing and serum metabolome by nuclear magnetic resonance (NMR) platform. The intensive weight loss period induced several metabolic adaptations, including a prominent suppression of transcriptomic signature for mitochondrial OXPHOS and ribosome biogenesis. The upstream regulator analysis suggested that this reprogramming of cellular energy metabolism may be mediated via AMPK/PGC1-α signaling and mTOR/eIF2 signaling-dependent pathways. Our findings show for the first time that prolonged energy deprivation induced modulation of mitochondrial metabolism can be observed through minimally invasive measures of leukocyte transcriptome and serum metabolome at systemic level, suggesting that adaptation to energy deficit is broader in humans than previously thought.

Design and synthesis of nature-inspired chromenopyrroles as potential modulators of mitochondrial metabolism

Chromenopyrrole derivatives with multiple stereocenters and variable ring fusion pattern are found in many natural products and biologically appealing molecules. By employing a build/couple/pair strategy, we have recently reported on the discovery of a serendipitous cascade to access a diverse collection of chromenopyrroles. This protocol features a one-pot cascade that includes the generation of azomethine ylide and intramolecular [3 + 2]-cycloaddition. Phenotypic screening of the developed pilot library enabled the identification of chemical probes that efficiently suppress mitochondrial membrane potential, elevate reactive oxygen species content, and deplete ATP content in a hepatoma cell line (Hepa1-6), without affecting the proliferation of T- or B-cells. This selective targeting represents a new approach for the treatment of cancer.

Mitochondria of transformed cell as a target of antitumor influence

Mitochondria are intracellular organelles in eukaryotic cells that participate in bioenergy metabolism and cell homeostasis, including ATP generation through electron transport and oxidative phosphorylation in combination with oxidation of metabolites by the tricarboxylic acid cycle and fatty acid catabolism via β-oxidation. the production of reactive oxygen species, as well as the initiation and implementation of apoptosis. Mitochondria play a crucial role in cellular energy metabolism and the regulation of programmed cell death. mitochondria activate numerous signaling pathways associated with cell death. Mitochondria have the ability to control the activation of programmed cell death by regulating the translocation of proapoptotic proteins from the intermediate space of mitochondria to the cytosol. This is the reason for the emergence of a new discipline — mitochondrial medicine. The review examined and analyzed scientific publications on the role of mitochondria in the life support of transformed cells, the study of their functioning and structurally functional dysfunctions, as part of mitochondrial medicine. Mitochondrial medicine is a developing discipline whose significance stems from the central function of mitochondria in the production of adenosine triphosphate, the generation of reactive oxygen species, and cell death due to necrosis or apoptosis. Consequently, mitochondrial dysfunction plays an important role in the pathophysiology of cancer, many other common diseases and side effects of drugs. Perhaps the combined use of modulators of mitochondrial metabolism and antitumor therapy will contribute to the emergence of a new direction in antitumor treatment, which will significantly increase the effectiveness of cancer treatment.

Preventive and Therapeutic Potential of Physical Exercise in Neurodegenerative Diseases.

Significance: The prevalence and incidence of age-related neurodegenerative diseases (NDDs) tend to increase along with the enhanced average of the world life expectancy. NDDs are a major cause of morbidity and disability, affecting the health care, social and economic systems with a significant impact. Critical Issues and Recent Advances: Despite the worldwide burden of NDDs and the ongoing research efforts to increase the underlying molecular mechanisms involved in NDD pathophysiologies, pharmacological therapies have been presenting merely narrow benefits. On the contrary, absent of detrimental side effects but growing merits, regular physical exercise (PE) has been considered a prone pleiotropic nonpharmacological alternative able to modulate brain structure and function, thereby stimulating a healthier and "fitness" neurological phenotype. Future Directions: This review summarizes the state of the art of some peripheral and central-related mechanisms that underlie the impact of PE on brain plasticity as well as its relevance for the prevention and/or treatment of NDDs. Nevertheless, further studies are needed to better clarify the molecular signaling pathways associated with muscle contractions-related myokines release and its plausible positive effects in the brain. In addition, particular focus of research should address the role of PE in the modulation of mitochondrial metabolism and oxidative stress in the context of NDDs.

Therapeutic Concentrations of Statins Hyperpolarize Mitochondria and Inhibit Cell Proliferation Without Promoting Cell Death in Hepatocarcinoma Cells

Statins, widely used inhibitors of HMG-CoA reductase to decrease cholesterol, may also decrease the risk of developing liver, breast, and lung cancer. Concentrations of statins (10-100 µM) typically used in vitro are 1,000- to 10,000-fold higher than plasmatic therapeutic concentrations (10-100 nM). In preliminary work, we showed that simvastatin (SIM) and lovastatin at 10 µM increased mitochondrial membrane potential (ΔΨ) and decreased respiration and ATP production in HepG2, Huh7 and HCC4006 cells. We also showed that the bcl-XL inhibitor ABT-737 prevents mitochondrial hyperpolarization induced by the ATP synthase inhibitor, oligomycin. Here, we hypothesize that SIM at nanomolar concentrations increases ΔΨ contributing to decreased cell proliferation and that both mitochondrial hyperpolarization and inhibition of cell proliferation are influenced by bcl-XL. Confocal fluorescence microscopy of tetramethylrhodamine methyl ester assessed ΔΨ. Cholesterol levels were determined using Amplex Red after lipid extraction from whole HepG2 cells and isolated mitochondria. Respiration was assessed using a Seahorse XFe96 Analyzer and cell proliferation by direct counting of live/dead cells. SIM (20 nM to 100 nM) increased mitochondrial ΔΨ in a dose-dependent fashion in HepG2 cells at 24 h. SIM (100 nM for 24 h and 10 µM for 6 h) increased ΔΨ by ∼120 % and 160 % respectively. High ΔΨ promoted by SIM (100 nM and 10 µM) was sustained for at least 72 h. ABT-737 (1 µM) blocked mitochondrial hyperpolarization induced by SIM. SIM at 100 nM and 10 µM decreased cell proliferation by ∼49% and 82% respectively without promoting cell killing. The inhibition of cell proliferation was blocked by pretreatment with ABT-737. Therapeutic concentrations of statins decreased cell proliferation without promoting cell killing possibly by modulation of mitochondrial metabolism mediated by bcl-XL.

Measurement of Mitochondrial Mass and Membrane Potential in Hematopoietic Stem Cells and T-cells by Flow Cytometry.

A fine balance of quiescence, self-renewal, and differentiation is key to preserve the hematopoietic stem cell (HSC) pool and maintain lifelong production of all mature blood cells. In recent years cellular metabolism has emerged as a crucial regulator of HSC function and fate. We have previously demonstrated that modulation of mitochondrial metabolism influences HSC fate. Specifically, by chemically uncoupling the electron transport chain we were able to maintain HSC function in culture conditions that normally induce rapid differentiation. However, limiting HSC numbers often precludes the use of standard assays to measure HSC metabolism and therefore predict their function. Here, we report a simple flow cytometry assay that allows reliable measurement of mitochondrial membrane potential and mitochondrial mass in scarce cells such as HSCs. We discuss the isolation of HSCs from mouse bone marrow and measurement of mitochondrial mass and membrane potential post ex vivo culture. As an example, we show the modulation of these parameters in HSCs via treatment with a metabolic modulator. Moreover, we extend the application of this methodology on human peripheral blood-derived T cells and human tumor infiltrating lymphocytes (TILs), showing dramatic differences in their mitochondrial profiles, possibly reflecting different T cell functionality. We believe this assay can be employed in screenings to identify modulators of mitochondrial metabolism in various cell types in different contexts.

P3487Pre-ischemic succinate dehydrogenase inhibition is protective against ischemia-reperfusion injury in rat and human myocardium with and without diabetes mellitus

Abstract Background Ischemia-reperfusion (IR) injury can be attenuated through modulation of mitochondrial metabolism with succinate dehydrogenase (SDH) inhibition by dimethyl malonate (DiMAL). However, it is unknown whether SDH inhibition yields protection in aged, diabetic individuals and whether protection translate into human cardiac tissue. Purpose We wished to evaluate if SDH inhibition can be protective in aged, diabetic rat hearts and could be translated to the human myocardium. Methods We studied infarct size in aged non-diabetic and diabetic rat hearts subjected to isolated, retrograde perfusion and global ischemia and reperfusion. The hearts were randomized to: Sham hearts, IR injured hearts and IR injured hearts co-perfused with 0.1 mM or 0.6 mM DiMAL. Infarct size and mitochondrial respiratory capacity were evaluated post-ischemic. To translate our findings into human cardiac tissue, we tested the efficacy of DiMAL treatment in human atrial trabeculae from patients with and without diabetes mellitus undergoing cardiac surgery. We randomized atrial trabeculae to: IR injury, IR injury treated with ischemic preconditioning (IPC) and IR injury treated with 5 mM DiMAL. Contractile force recovery and mitochondrial respiratory capacity were evaluated post-ischemic. Results In non-diabetic rat hearts, DiMAL 0.1 mM reduced infarct size/area-at-risk (IS/AAR) compared to IR hearts (53±7% vs. 69±6% of IS/AAR; p<0.05). In diabetic hearts, an increased concentration of 0.6 mM DiMAL was required to confer protection (62±13% vs. 80±8% of IS/AAR; p<0.05). Mitochondrial complex I linked respiratory capacity was higher in both Sham and DiMAL 0.1 mM groups than with DiMAL 0.6 mM in non-diabetic hearts (157.4±26.5 and 140.0±35.3 vs. 81.8±42.4 pmol O2/(s*mg); p<0.01 and p<0.05). In trabeculae from humans without diabetes mellitus, IPC and DiMAL improved contractile force recovery compared to IR (43±12% and 43±13% vs. 23±13%, p<0.05). In trabeculae from patients with diabetes, IPC but not DiMAL administration improved contractile force recovery compared to IR (48±16% and 19±7% vs. 25±13%, p<0.01 and p>0.99). Mitochondrial complex I linked respiration capacity did not improve by IPC or DiMAL compared to IR in non-diabetic trabeculae (p>0.99). Conclusion Inhibition of the SDH by DiMAL protects the aged non-diabetic and diabetic heart from IR injury but the threshold for cardioprotection is increased in diabetic rat hearts. DiMAL provides cardioprotection similar to IPC in non-diabetic trabeculae but does not protect the human diabetic myocardium when given in identical concentrations. SDH inhibition can provide protection in both animal and human tissue and may provide a new target for pharmacologic conditioning in patients.

Mitochondrial targets of metformin-Are they physiologically relevant?

Metformin is the most widely prescribed treatment of hyperglycemia and type II diabetes since 1970s. During the last 15 years, its popularity increased due to epidemiological evidence, that metformin administration reduces incidence of cancer. However, despite the ongoing effort of many researchers, the molecular mechanisms underlying antihyperglycemic or antineoplastic action of metformin remain elusive. Most frequently, metformin is associated with modulation of mitochondrial metabolism leading to lowering of blood glucose or activation of antitumorigenic pathways. Here we review the reported effects of metformin on mitochondrial metabolism and their potential relevance as effective molecular targets with beneficial therapeutic outcome.

Statin‐dependent modulation of mitochondrial metabolism in cancer cells is independent of cholesterol content

Statins, widely used to treat hypercholesterolemia, inhibit the 3-hydroxy-3-methylglutaryl-coenzyme A reductase, the rate-limiting enzyme of de novo cholesterol (Chol) synthesis. Statins have been also reported to slow tumor progression. In cancer cells, ATP is generated both by glycolysis and oxidative phosphorylation. Mitochondrial membrane potential (ΔΨ), a readout of mitochondrial metabolism, is sustained by the oxidation of respiratory substrates in the Krebs cycle to generate NADH and flavin adenine dinucleotide, which are further oxidized by the respiratory chain. Here, we studied the short-term effects of statins (3-24 h) on mitochondrial metabolism on cancer cells. Lovastatin (LOV) and simvastatin (SIM) increased ΔΨ in HepG2 and Huh7 human hepatocarcinoma cells and HCC4006 human lung adenocarcinoma cells. Mitochondrial hyperpolarization after LOV and SIM was dose and time dependent. Maximal increase in ΔΨ occurred at 10 µM and 24 h for both statins. The structurally unrelated atorvastatin also hyperpolarized mitochondria in HepG2 cells. Cellular and mitochondrial Chol remained unchanged after SIM. Both LOV and SIM decreased basal respiration, ATP-linked respiration, and ATP production. LOV and SIM did not change the rate of lactic acid production. In summary, statins modulate mitochondrial metabolism in cancer cells independently of the Chol content in cellular membranes without affecting glycolysis.-Christie, C. F., Fang, D., Hunt, E. G., Morris, M. E., Rovini, A., Heslop, K. A., Beeson, G. C., Beeson, C. C., Maldonado, E. N. Statin-dependent modulation of mitochondrial metabolism in cancer cells is independent of cholesterol content.

Modulation of mitochondrial metabolism as a biochemical trait in blood feeding organisms: the redox vampire hypothesis redux

Hematophagous organisms undergo remarkable metabolic changes during the blood digestion process, increasing fermentative glucose metabolism, and reducing respiratory rates, both consequence of functional mitochondrial remodeling. Here, we review the pathways involved in energy metabolism and mitochondrial functionality in a comparative framework across different hematophagous species, and consider how these processes regulate redox homeostasis during blood digestion. The trend across distinct species indicate that a switch in energy metabolism might represent an important defensive mechanism to avoid the potential harmful interaction of oxidants generated from aerobic energy metabolism with products derived from blood digestion. Indeed, in insect vectors, blood feeding transiently reduces respiratory rates and oxidant production, irrespective of tissue and insect model. On the other hand, a different scenario is observed in several unrelated parasite species when exposed to blood digestion products, as respiratory rates reduce and mitochondrial oxidant production increase. The emerging picture indicates that re-wiring of energy metabolism, through reduced mitochondrial function, culminates in improved tolerance to redox insults and seems to represent a key step for hematophagous organisms to cope with the overwhelming and potentially toxic blood meal.

Amyloid precursor protein drives down-regulation of mitochondrial oxidative phosphorylation independent of amyloid beta

Amyloid precursor protein (APP) and its extracellular domain, soluble APP alpha (sAPPα) play important physiological and neuroprotective roles. However, rare forms of familial Alzheimer’s disease are associated with mutations in APP that increase toxic amyloidogenic cleavage of APP and produce amyloid beta (Aβ) at the expense of sAPPα and other non-amyloidogenic fragments. Although mitochondrial dysfunction has become an established hallmark of neurotoxicity, the link between Aβ and mitochondrial function is unclear. In this study we investigated the effects of increased levels of neuronal APP or Aβ on mitochondrial metabolism and gene expression, in human SH-SY5Y neuroblastoma cells. Increased non-amyloidogenic processing of APP, but not Aβ, profoundly decreased respiration and enhanced glycolysis, while mitochondrial DNA (mtDNA) transcripts were decreased, without detrimental effects to cell growth. These effects cannot be ascribed to Aβ toxicity, since higher levels of endogenous Aβ in our models do not cause oxidative phosphorylation (OXPHOS) perturbations. Similarly, chemical inhibition of β-secretase decreased mitochondrial respiration, suggesting that non-amyloidogenic processing of APP may be responsible for mitochondrial changes. Our results have two important implications, the need for caution in the interpretation of mitochondrial perturbations in models where APP is overexpressed, and a potential role of sAPPα or other non-amyloid APP fragments as acute modulators of mitochondrial metabolism.

MicroRNA-17 family promotes polycystic kidney disease progression through modulation of mitochondrial metabolism

Autosomal dominant polycystic kidney disease (ADPKD) is the most frequent genetic cause of renal failure. Here we identify miR-17 as a target for the treatment of ADPKD. We report that miR-17 is induced in kidney cysts of mouse and human ADPKD. Genetic deletion of the miR-17∼92 cluster inhibits cyst proliferation and PKD progression in four orthologous, including two long-lived, mouse models of ADPKD. Anti-miR-17 treatment attenuates cyst growth in short-term and long-term PKD mouse models. miR-17 inhibition also suppresses proliferation and cyst growth of primary ADPKD cysts cultures derived from multiple human donors. Mechanistically, c-Myc upregulates miR-17∼92 in cystic kidneys, which in turn aggravates cyst growth by inhibiting oxidative phosphorylation and stimulating proliferation through direct repression of Pparα. Thus, miR-17 family is a promising drug target for ADPKD, and miR-17-mediated inhibition of mitochondrial metabolism represents a potential new mechanism for ADPKD progression.

Cross-talk between branched-chain amino acids and hepatic mitochondria is compromised in nonalcoholic fatty liver disease

Elevated plasma branched-chain amino acids (BCAA) in the setting of insulin resistance have been relevant in predicting type 2 diabetes mellitus (T2DM) onset, but their role in the etiology of hepatic insulin resistance remains uncertain. We determined the link between BCAA and dysfunctional hepatic tricarboxylic acid (TCA) cycle, which is a central feature of hepatic insulin resistance and nonalcoholic fatty liver disease (NAFLD). Plasma metabolites under basal fasting and euglycemic hyperinsulinemic clamps (insulin stimulation) were measured in 94 human subjects with varying degrees of insulin sensitivity to identify their relationships with insulin resistance. Furthermore, the impact of elevated BCAA on hepatic TCA cycle was determined in a diet-induced mouse model of NAFLD, utilizing targeted metabolomics and nuclear magnetic resonance (NMR)-based metabolic flux analysis. Insulin stimulation revealed robust relationships between human plasma BCAA and indices of insulin resistance, indicating chronic metabolic overload from BCAA. Human plasma BCAA and long-chain acylcarnitines also showed a positive correlation, suggesting modulation of mitochondrial metabolism by BCAA. Concurrently, mice with NAFLD failed to optimally induce hepatic mTORC1, plasma ketones, and hepatic long-chain acylcarnitines, following acute elevation of plasma BCAA. Furthermore, elevated BCAA failed to induce multiple fluxes through hepatic TCA cycle in mice with NAFLD. Our data suggest that BCAA are essential to mediate efficient channeling of carbon substrates for oxidation through mitochondrial TCA cycle. Impairment of BCAA-mediated upregulation of the TCA cycle could be a significant contributor to mitochondrial dysfunction in NAFLD.